For complete and up to date information on this and other items, consult my Curriculum vitae (updated, 12/16), and website; URL:

Academic Appointments

Administrative Appointments

  • Member, Stanford Panel on human subjects, Stanford University (2000 - 2004)
  • Member, Stanford Recombinant DNA Panel, Stanford University (1979 - 1982)
  • Chair, Stanford Recombiant DNA Panel, Stanford University (1981 - 1982)
  • Chair, Department Admissions Committee, Stanford University (1985 - 1988)
  • Member, Administrative Panel on Biosafety, Stanford University (1995 - 1996)
  • Member, Stanford Human Subjects Panel (2000 - 2004)
  • Senator, Medical School senate (2006 - 2012)
  • Member, MS senate steering committee (2008 - 2012)
  • Chair, MS senate task force on posdoctoral affairs (2009 - 2012)

Honors & Awards

  • Editorial Board, Annual Review of Microbiology (1980-1983, 2005)
  • Member Study Section, National Institutes of Health (2003)
  • Member Study Section, Department of Energy (1996, 1997, 2002)
  • Editorial Board Member: Journal of Bacteriology:, American Society for Microbiology (1987-1993)
  • Member Study Section, NIH Environmental Institute (2003)
  • Member Study Section, National Aeronautics and Space Administration (2001, 2004, 2008, 2011)
  • Board Member, Scientific Advisory Board, Institute of Molecular Medicine, New York, Kolkata (2002-2004)
  • Member, Advisory Board, Chembiotek (2002 - 2004)
  • Member, Advisory Board, Chemgen Pharma International (2004-2008)
  • Expert consultant and witness, Law Offices of Swidler Berlin Shereff Friedman, LLP. (1999 - 2001)
  • Fulbright Scholar, Fulbright Foundation (1964-1971)
  • Foundation for Microbiology Lecturer, American Society for Microbiology (1991-1993)
  • Review Committee Member, Accreditation Board for Engineering and Technology (1992)
  • Star Award, Environmental Protection Agency (1991,1997)
  • Elected Fellow, American Academy of Microbiology (1994-)
  • Chartered Member, Drug Discovery & Molecular Pharmacology Study section, NCI (2008-2012)
  • Editor-in-Chief, Open Journal of Applied Sciences (2012)
  • Editorial Board, Cancer Management and Research (2008-present)
  • Editorial Board, Journal of Molecular Imaging & Dynamic (2010-present)

Boards, Advisory Committees, Professional Organizations

  • Listed in, Wikipedia (2021 - Present)

Professional Education

  • Ph. D., University of California, Microbiology (1969)

Community and International Work

  • Exosome-mediated specific therapy of cancer


    Making targeted exosomes

    Partnering Organization(s)

    NIH EERC consortium; several Stanford faculty

    Populations Served




    Ongoing Project


    Opportunities for Student Involvement


  • Bacterial antibiotic resistance in space flight, Stanford University; NASA Ames


    Genetic basis of resistance

    Partnering Organization(s)



    Bay Area

    Ongoing Project


    Opportunities for Student Involvement


  • Nuclear waste remediation



    Partnering Organization(s)

    Cadarache, France



    Ongoing Project


    Opportunities for Student Involvement


  • Lectures

    Ongoing Project


    Opportunities for Student Involvement



  • AC Matin. "United StatesFollow my website; URL hyperlinked in, "LINKS""

Current Research and Scholarly Interests

Please follow myResearch/
Lab website link:

2023-24 Courses

All Publications

  • Medical Use of mRNA-Based Directed Gene Delivery MESSENGER RNA THERAPEUTICS Matin, A. C., Forterre, A., Jurga, S., Barciszewski, J. 2022; 13: 93-112
  • Response to Comments on "EcAMSat spaceflight measurements of the role of sigmas in antibiotic resistance of stationary phase Escherichia coli in microgravity". Life sciences in space research Padgen, M. R., Parra, M. P., Ricco, A. J., Matin, A. C. 2021; 29: 85–86

    View details for DOI 10.1016/j.lssr.2021.03.006

    View details for PubMedID 33888293

  • Extracellular vesicle-mediated in vitro transcribed mRNA delivery for treatment of HER2+ breast cancer xenografts in mice by prodrug CB1954 without general toxicity. Molecular cancer therapeutics Forterre, A. V., Wang, J. H., Delcayre, A. n., Kim, K. n., Green, C. n., Pegram, M. D., Jeffrey, S. S., Matin, A. C. 2020


    Prodrugs are harmless until activated by a bacterial or viral gene product; they constitute the basis of gene delivered prodrug therapies called GDEPTs, which can kill tumors without major side effects. Previously, we utilized the prodrug CNOB (not clinically tested) and enzyme HChrR6 in GDEPT to generate the drug MCHB in tumors. Extracellular vesicles (EVs) were used for directed gene delivery and HChrR6 mRNA as gene. Here, the clinical transfer of this approach is enhanced by: 1. Use of CB1954 (tretazicar) for which safe human dose is established; HChrR6 can activate this prodrug. 2. EVs delivered in vitro transcribed (IVT) HChrR6 mRNA, eliminating the potentially harmful plasmid transfection of EV-producer cells we utilized previously; this has not been done before. IVT mRNA loading of EVs required several steps. Naked mRNA being unstable, we ensured its prodrug activating functionality at each step. This was not possible using tretazicar itself; we relied instead on HChrR6's ability to convert CNOB into MCHB, whose fluorescence is easily visualizable. HChrR6 mRNA-translated product's ability to generate fluorescence from CNOB vicariously indicated its competence for tretazicar activation. 3. Systemic IVT mRNA loaded EVs displaying an anti-HER2 scFv ("IVT EXO-DEPTs") and tretazicar caused growth arrest of human HER2+ breast cancer xenografts in athymic mice. As this occurred without injury to other tissues, absence of off-target mRNA delivery is strongly indicated. Many cancer sites are not amenable for direct gene injection, but current GDEPTs require this. In circumventing this need, a major advance in GDEPT applicability has been accomplished.

    View details for DOI 10.1158/1535-7163.MCT-19-0928

    View details for PubMedID 31941722

  • EcAMSat spaceflight measurements of the role of σs in antibiotic resistance of stationary phase Escherichia coli in microgravity. Life sciences in space research Padgen, M. R., Lera, M. P., Parra, M. P., Ricco, A. J., Chin, M. n., Chinn, T. N., Cohen, A. n., Friedericks, C. R., Henschke, M. B., Snyder, T. V., Spremo, S. M., Wang, J. H., Matin, A. C. 2020; 24: 18–24


    We report the results of the EcAMSat (Escherichia coli Antimicrobial Satellite) autonomous space flight experiment, investigating the role of σs in the development of antibiotic resistance in uropathogenic E. coli (UPEC) in microgravity (µ-g). The presence of σs, encoded by the rpoS gene, has been shown to increase antibiotic resistance in Earth gravity, but it was unknown if this effect occurs in µ-g. Two strains, wildtype (WT) UPEC and its isogenic ΔrpoS mutant, were grown to stationary phase aboard EcAMSat, an 11-kg small satellite, and in a parallel ground-based control experiment; cell growth rates for the two strains were found to be unaltered by µ-g. After starvation for over 24 h, stationary-phase cells were incubated with three doses of gentamicin (Gm), a common treatment for urinary tract infections (which have been reported in astronauts). Cellular metabolic activity was measured optically using the redox-based indicator alamarBlue (aB): both strains exhibited slower metabolism in µ-g, consistent with results from previous smallsat missions. The results also showed that µ-g did not enhance UPEC resistance to Gm; in fact, both strains were more susceptible to Gm in µ-g. It was also found, via a second ground-control experiment, that multi-week storage in the payload hardware stressed the cells, potentially obscuring small differential effects of the antibiotic between WT and mutant and/or between µ-g and ground. Overall, results showed that the ∆rpoS mutant was 34-37% less metabolically active than the WT for four different sets of conditions: ground without Gm, ground with Gm; µ-g without Gm, µ-g with Gm. We conclude therefore that the rpoS gene and its downstream products are important therapeutic targets for treating bacterial infections in space, much as they are on the ground.

    View details for DOI 10.1016/j.lssr.2019.10.007

    View details for PubMedID 31987476

  • Phenotyping antibiotic resistance with single-cell resolution for the detection of heteroresistance SENSORS AND ACTUATORS B-CHEMICAL Lyu, F., Pan, M., Patil, S., Wang, J., Matin, A. C., Andrews, J. R., Tang, S. Y. 2018; 270: 396–404
  • Anti-HER2 scFv-directed extracellular vesicle-mediated mRNA-based gene delivery inhibits growth of HER2-positive human breast tumor xenografts by prodrug activation. Molecular cancer therapeutics Wang, J., Forterre, A. V., Zhao, J., Frimannsson, D. O., Delcayre, A., Antes, T. J., Efron, B., Jeffrey, S. S., Pegram, M. D., Matin, A. C. 2018


    This paper deals with specific targeting of the prodrug/enzyme regimen, CNOB/HChrR6, to treat a serious disease namely HER2+ve human breast cancer with minimal off-target toxicity. HChrR6 is an improved bacterial enzyme that converts CNOB into the cytotoxic drug MCHB. Extracellular vesicles (EVs) were used for mRNA-based HchrR6 gene delivery: EVs may cause minimal immune rejection, and mRNA may be superior to DNA for gene delivery. To confine HChrR6 generation and CNOB activation to the cancer, the EVHB chimeric protein was constructed. It contains high affinity anti-HER2 scFv antibody (ML39) and is capable of latching on to EV surface. Cells transfected with EVHB-encoding plasmid generated EVs displaying this protein ("directed EVs"). Transfection of a separate batch of cells with the new plasmid, XPort/HChrR6, generated EVs containing HChrR6 mRNA; incubation with pure EVHB enabled these to target the HER2 receptor, generating "EXO-DEPT" EVs. EXO-DEPT treatment specifically enabled HER2-overexpressing BT474 cells to convert CNOB into MCHB in actinomycin D independent manner, showing successful and specific delivery of HCHrR6 mRNA. EXO-DEPTs --but not undirected EVs-- plus CNOB caused near-complete growth-arrest of orthotopic BT474 xenografts in vivo, demonstrating for the first time EV-mediated delivery of functional exogenous mRNA to tumors. EXO-DEPTs may be generated from patient's own dendritic cells to evade immune rejection, and without plasmids and their potentially harmful genetic material, raising the prospect of clinical use of this regimen. This approach can be employed to treat any disease overexpressing a specific marker.

    View details for PubMedID 29483213

  • Payload hardware and experimental protocol development to enable future testing of the effect of space microgravity on the resistance to gentamicin of uropathogenic Escherichia coli and its sigma(s)-deficient mutant LIFE SCIENCES IN SPACE RESEARCH Matin, A. C., Wang, J., Keyhan, M., Singh, R., Benoit, M., Parra, M. P., Padgen, M. R., Ricco, A. J., Chin, M., Friedericks, C. R., Chinn, T. N., Cohen, A., Henschke, M. B., Snyder, T. V., Lera, M. P., Ross, S. S., Mayberry, C. M., Choi, S., Wu, D. T., Tan, M. X., Boone, T. D., Beasley, C. C., Piccini, M. E., Spremo, S. M. 2017; 15: 1–10


    Human immune response is compromised and bacteria can become more antibiotic resistant in space microgravity (MG). We report that under low-shear modeled microgravity (LSMMG), stationary-phase uropathogenic Escherichia coli (UPEC) become more resistant to gentamicin (Gm), and that this increase is dependent on the presence of σs (a transcription regulator encoded by the rpoS gene). UPEC causes urinary tract infections (UTIs), reported to afflict astronauts; Gm is a standard treatment, so these findings could impact astronaut health. Because LSMMG findings can differ from MG, we report preparations to examine UPEC's Gm sensitivity during spaceflight using the E. coli Anti-Microbial Satellite (EcAMSat) as a free-flying "nanosatellite" in low Earth orbit. Within EcAMSat's payload, a 48-microwell fluidic card contains and supports study of bacterial cultures at constant temperature; optical absorbance changes in cell suspensions are made at three wavelengths for each microwell and a fluid-delivery system provides growth medium and predefined Gm concentrations. Performance characterization is reported here for spaceflight prototypes of this payload system. Using conventional microtiter plates, we show that Alamar Blue (AB) absorbance changes can assess the Gm effect on E. coli viability, permitting telemetric transfer of the spaceflight data to Earth. Laboratory results using payload prototypes are consistent with wellplate and flask findings of differential sensitivity of UPEC and its ∆rpoS strain to Gm. if σs plays the same role in space MG as in LSMMG and Earth gravity, countermeasures discovered in recent Earth studies (aimed at weakening the UPEC antioxidant defense) to control UPEC infections would prove useful also in space flights. Further, EcAMSat results should clarify inconsistencies from previous space experiments on bacterial antibiotic sensitivity and other issues.

    View details for DOI 10.1016/j.lssr.2017.05.001

    View details for Web of Science ID 000416873500001

    View details for PubMedID 29198308

  • Utilizing native fluorescence imaging, modeling and simulation to examine pharmacokinetics and therapeutic regimen of a novel anticancer prodrug. BMC cancer Wang, J., Endsley, A. N., Green, C. E., Matin, A. C. 2016; 16: 524-?


    Success of cancer prodrugs relying on a foreign gene requires specific delivery of the gene to the cancer, and improvements such as higher level gene transfer and expression. Attaining these objectives will be facilitated in preclinical studies using our newly discovered CNOB-GDEPT, consisting of the produrg: 6-chloro-9-nitro-5-oxo-5H-benzo-(a)-phenoxazine (CNOB) and its activating enzyme ChrR6, which generates the cytotoxic product 9-amino-6-chloro-5H-benzo[a]phenoxazine-5-one (MCHB). MCHB is fluorescent and can be noninvasively imaged in mice, and here we investigated whether MCHB fluorescence quantitatively reflects its concentration, as this would enhance its reporter value in further development of the CNOB-GDEPT therapeutic regimen. PK parameters were estimated and used to predict more effective CNOB administration schedules.CNOB (3.3 mg/kg) was injected iv in mice implanted with humanized ChrR6 (HChrR6)-expressing 4T1 tumors. Fluorescence was imaged in live mice using IVIS Spectrum, and quantified by Living Image 3.2 software. MCHB and CNOB were quantified also by LC/MS/MS analysis. We used non-compartmental model to estimate PK parameters. Phoenix WinNonlin software was used for simulations to predict a more effective CNOB dosage regimen.CNOB administration significantly prolonged mice survival. MCHB fluorescence quantitatively reflected its exposure levels to the tumor and the plasma, as verified by LC/MS/MS analysis at various time points, including at a low concentration of 2 ng/g tumor. The LC/MS/MS data were used to estimate peak plasma concentrations, exposure (AUC0-24), volume of distribution, clearance and half-life in plasma and the tumor. Simulations suggested that the CNOB-GDEPT can be a successful therapy without large increases in the prodrug dosage.MCHB fluorescence quantifies this drug, and CNOB can be effective at relatively low doses. MCHB fluorescence characteristics will expedite further development of CNOB-GDEPT by, for example, facilitating specific gene delivery to the tumor, its prolonged expression, as well as other attributes necessary for successful gene-delivered enzyme prodrug therapy.

    View details for DOI 10.1186/s12885-016-2508-6

    View details for PubMedID 27457630

    View details for PubMedCentralID PMC4960810

  • Differential fates of biomolecules delivered to target cells via extracellular vesicles. Proceedings of the National Academy of Sciences of the United States of America Kanada, M., Bachmann, M. H., Hardy, J. W., Frimannson, D. O., Bronsart, L., Wang, A., Sylvester, M. D., Schmidt, T. L., Kaspar, R. L., Butte, M. J., Matin, A. C., Contag, C. H. 2015; 112 (12): E1433-42


    Extracellular vesicles (EVs), specifically exosomes and microvesicles (MVs), are presumed to play key roles in cell-cell communication via transfer of biomolecules between cells. The biogenesis of these two types of EVs differs as they originate from either the endosomal (exosomes) or plasma (MVs) membranes. To elucidate the primary means through which EVs mediate intercellular communication, we characterized their ability to encapsulate and deliver different types of macromolecules from transiently transfected cells. Both EV types encapsulated reporter proteins and mRNA but only MVs transferred the reporter function to recipient cells. De novo reporter protein expression in recipient cells resulted only from plasmid DNA (pDNA) after delivery via MVs. Reporter mRNA was delivered to recipient cells by both EV types, but was rapidly degraded without being translated. MVs also mediated delivery of functional pDNA encoding Cre recombinase in vivo to tissues in transgenic Cre-lox reporter mice. Within the parameters of this study, MVs delivered functional pDNA, but not RNA, whereas exosomes from the same source did not deliver functional nucleic acids. These results have significant implications for understanding the role of EVs in cellular communication and for development of EVs as delivery tools. Moreover, studies using EVs from transiently transfected cells may be confounded by a predominance of pDNA transfer.

    View details for DOI 10.1073/pnas.1418401112

    View details for PubMedID 25713383

    View details for PubMedCentralID PMC4378439

  • Differential fates of biomolecules delivered to target cells via extracellular vesicles PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA Kanada, M., Bachmann, M. H., Hardy, J. W., Frimannson, D. O., Bronsart, L., Wang, A., Sylvester, M. D., Schmidt, T. L., Kaspar, R. L., Butte, M. J., Matin, A. C., Contag, C. H. 2015; 112 (12): E1433-E1442


    Extracellular vesicles (EVs), specifically exosomes and microvesicles (MVs), are presumed to play key roles in cell-cell communication via transfer of biomolecules between cells. The biogenesis of these two types of EVs differs as they originate from either the endosomal (exosomes) or plasma (MVs) membranes. To elucidate the primary means through which EVs mediate intercellular communication, we characterized their ability to encapsulate and deliver different types of macromolecules from transiently transfected cells. Both EV types encapsulated reporter proteins and mRNA but only MVs transferred the reporter function to recipient cells. De novo reporter protein expression in recipient cells resulted only from plasmid DNA (pDNA) after delivery via MVs. Reporter mRNA was delivered to recipient cells by both EV types, but was rapidly degraded without being translated. MVs also mediated delivery of functional pDNA encoding Cre recombinase in vivo to tissues in transgenic Cre-lox reporter mice. Within the parameters of this study, MVs delivered functional pDNA, but not RNA, whereas exosomes from the same source did not deliver functional nucleic acids. These results have significant implications for understanding the role of EVs in cellular communication and for development of EVs as delivery tools. Moreover, studies using EVs from transiently transfected cells may be confounded by a predominance of pDNA transfer.

    View details for DOI 10.1073/pnas.1418401112

    View details for Web of Science ID 000351477000008

    View details for PubMedID 25713383

    View details for PubMedCentralID PMC4378439

  • Sigma S-Dependent Antioxidant Defense Protects Stationary-Phase Escherichia coli against the Bactericidal Antibiotic Gentamicin ANTIMICROBIAL AGENTS AND CHEMOTHERAPY Wang, J., Singh, R., Benoit, M., Keyhan, M., Sylvester, M., Hsieh, M., Thathireddy, A., Hsieh, Y., Matin, A. C. 2014; 58 (10): 5964-5975


    Stationary-phase bacteria are important in disease. The σ(s)-regulated general stress response helps them become resistant to disinfectants, but the role of σ(s) in bacterial antibiotic resistance has not been elucidated. Loss of σ(s) rendered stationary-phase Escherichia coli more sensitive to the bactericidal antibiotic gentamicin (Gm), and proteomic analysis suggested involvement of a weakened antioxidant defense. Use of the psfiA genetic reporter, 3'-(p-hydroxyphenyl) fluorescein (HPF) dye, and Amplex Red showed that Gm generated more reactive oxygen species (ROS) in the mutant. HPF measurements can be distorted by cell elongation, but Gm did not affect stationary-phase cell dimensions. Coadministration of the antioxidant N-acetyl cysteine (NAC) decreased drug lethality particularly in the mutant, as did Gm treatment under anaerobic conditions that prevent ROS formation. Greater oxidative stress, due to insufficient quenching of endogenous ROS and/or respiration-linked electron leakage, therefore contributed to the greater sensitivity of the mutant; infection by a uropathogenic strain in mice showed this to be the case also in vivo. Disruption of antioxidant defense by eliminating the quencher proteins, SodA/SodB and KatE/SodA, or the pentose phosphate pathway proteins, Zwf/Gnd and TalA, which provide NADPH for ROS decomposition, also generated greater oxidative stress and killing by Gm. Thus, besides its established mode of action, Gm also kills stationary-phase bacteria by generating oxidative stress, and targeting the antioxidant defense of E. coli can enhance its efficacy. Relevant aspects of the current controversy on the role of ROS in killing by bactericidal drugs of exponential-phase bacteria, which represent a different physiological state, are discussed.

    View details for DOI 10.1128/AAC.03683-14

    View details for Web of Science ID 000344157500040

    View details for PubMedCentralID PMC4187989

  • Sigma S-dependent antioxidant defense protects stationary-phase Escherichia coli against the bactericidal antibiotic gentamicin. Antimicrobial agents and chemotherapy Wang, J., Singh, R., Benoit, M., Keyhan, M., Sylvester, M., Hsieh, M., Thathireddy, A., Hsieh, Y., Matin, A. C. 2014; 58 (10): 5964-5975


    Stationary-phase bacteria are important in disease. The σ(s)-regulated general stress response helps them become resistant to disinfectants, but the role of σ(s) in bacterial antibiotic resistance has not been elucidated. Loss of σ(s) rendered stationary-phase Escherichia coli more sensitive to the bactericidal antibiotic gentamicin (Gm), and proteomic analysis suggested involvement of a weakened antioxidant defense. Use of the psfiA genetic reporter, 3'-(p-hydroxyphenyl) fluorescein (HPF) dye, and Amplex Red showed that Gm generated more reactive oxygen species (ROS) in the mutant. HPF measurements can be distorted by cell elongation, but Gm did not affect stationary-phase cell dimensions. Coadministration of the antioxidant N-acetyl cysteine (NAC) decreased drug lethality particularly in the mutant, as did Gm treatment under anaerobic conditions that prevent ROS formation. Greater oxidative stress, due to insufficient quenching of endogenous ROS and/or respiration-linked electron leakage, therefore contributed to the greater sensitivity of the mutant; infection by a uropathogenic strain in mice showed this to be the case also in vivo. Disruption of antioxidant defense by eliminating the quencher proteins, SodA/SodB and KatE/SodA, or the pentose phosphate pathway proteins, Zwf/Gnd and TalA, which provide NADPH for ROS decomposition, also generated greater oxidative stress and killing by Gm. Thus, besides its established mode of action, Gm also kills stationary-phase bacteria by generating oxidative stress, and targeting the antioxidant defense of E. coli can enhance its efficacy. Relevant aspects of the current controversy on the role of ROS in killing by bactericidal drugs of exponential-phase bacteria, which represent a different physiological state, are discussed.

    View details for DOI 10.1128/AAC.03683-14

    View details for PubMedID 25070093

  • Microgravity Alters the Physiological Characteristics of Escherichia coli O157:H7 ATCC 35150, ATCC 43889, and ATCC 43895 under Different Nutrient Conditions APPLIED AND ENVIRONMENTAL MICROBIOLOGY KIM, H. W., Matin, A., Rhee, M. S. 2014; 80 (7): 2270-2278


    The aim of this study is to provide understanding of microgravity effects on important food-borne bacteria, Escherichia coli O157:H7 ATCC 35150, ATCC 43889, and ATCC 43895, cultured in nutrient-rich or minimal medium. Physiological characteristics, such as growth (measured by optical density and plating), cell morphology, and pH, were monitored under low-shear modeled microgravity (LSMMG; space conditions) and normal gravity (NG; Earth conditions). In nutrient-rich medium, all strains except ATCC 35150 showed significantly higher optical density after 6 h of culture under LSMMG conditions than under NG conditions (P < 0.05). LSMMG-cultured cells were approximately 1.8 times larger than NG-cultured cells at 24 h; therefore, it was assumed that the increase in optical density was due to the size of individual cells rather than an increase in the cell population. The higher pH of the NG cultures relative to that of the LSMMG cultures suggests that nitrogen metabolism was slower in the latter. After 24 h of culturing in minimal media, LSMMG-cultured cells had an optical density 1.3 times higher than that of NG-cultured cells; thus, the higher optical density in the LSMMG cultures may be due to an increase in both cell size and number. Since bacteria actively grew under LSMMG conditions in minimal medium despite the lower pH, it is of some concern that LSMMG-cultured E. coli O157:H7 may be able to adapt well to acidic environments. These changes may be caused by changes in nutrient metabolism under LSMMG conditions, although this needs to be demonstrated in future studies.

    View details for DOI 10.1128/AEM.04037-13

    View details for Web of Science ID 000332840700026

    View details for PubMedID 24487539

  • Patient-derived xenografts of triple-negative breast cancer reproduce molecular features of patient tumors and respond to mTOR inhibition BREAST CANCER RESEARCH Zhang, H., Cohen, A. L., Krishnakumar, S., Wapnir, I. L., Veeriah, S., Deng, G., Coram, M. A., Piskun, C. M., Longacre, T. A., Herrler, M., Frimannsson, D. O., Telli, M. L., Dirbas, F. M., Matin, A. C., Dairkee, S. H., Larijani, B., Glinsky, G. V., Bild, A. H., Jeffrey, S. S. 2014; 16 (2)

    View details for DOI 10.1186/bcr3640

    View details for Web of Science ID 000338990900021

  • Crystal Structure of ChrR-A Quinone Reductase with the Capacity to Reduce Chromate PLOS ONE Eswaramoorthy, S., Poulain, S., Hienerwadel, R., Bremond, N., Sylvester, M. D., Zhang, Y., Berthomieu, C., van der Lelie, D., Matin, A. 2012; 7 (4)


    The Escherichia coli ChrR enzyme is an obligatory two-electron quinone reductase that has many applications, such as in chromate bioremediation. Its crystal structure, solved at 2.2 Å resolution, shows that it belongs to the flavodoxin superfamily in which flavin mononucleotide (FMN) is firmly anchored to the protein. ChrR crystallized as a tetramer, and size exclusion chromatography showed that this is the oligomeric form that catalyzes chromate reduction. Within the tetramer, the dimers interact by a pair of two hydrogen bond networks, each involving Tyr128 and Glu146 of one dimer and Arg125 and Tyr85 of the other; the latter extends to one of the redox FMN cofactors. Changes in each of these amino acids enhanced chromate reductase activity of the enzyme, showing that this network is centrally involved in chromate reduction.

    View details for DOI 10.1371/journal.pone.0036017

    View details for Web of Science ID 000305336000101

    View details for PubMedID 22558308

    View details for PubMedCentralID PMC3338774

  • New Device for High-Throughput Viability Screening of Flow Biofilms APPLIED AND ENVIRONMENTAL MICROBIOLOGY Benoit, M. R., Conant, C. G., Ionescu-Zanetti, C., Schwartz, M., Matin, A. 2010; 76 (13): 4136-4142


    Control of biofilms requires rapid methods to identify compounds effective against them and to isolate resistance-compromised mutants for identifying genes involved in enhanced biofilm resistance. While rapid screening methods for microtiter plate well ("static") biofilms are available, there are no methods for such screening of continuous flow biofilms ("flow biofilms"). Since the latter biofilms more closely approximate natural biofilms, development of a high-throughput (HTP) method for screening them is desirable. We describe here a new method using a device comprised of microfluidic channels and a distributed pneumatic pump (BioFlux) that provides fluid flow to 96 individual biofilms. This device allows fine control of continuous or intermittent fluid flow over a broad range of flow rates, and the use of a standard well plate format provides compatibility with plate readers. We show that use of green fluorescent protein (GFP)-expressing bacteria, staining with propidium iodide, and measurement of fluorescence with a plate reader permit rapid and accurate determination of biofilm viability. The biofilm viability measured with the plate reader agreed with that determined using plate counts, as well as with the results of fluorescence microscope image analysis. Using BioFlux and the plate reader, we were able to rapidly screen the effects of several antimicrobials on the viability of Pseudomonas aeruginosa PAO1 flow biofilms.

    View details for DOI 10.1128/AEM.03065-09

    View details for Web of Science ID 000279082800003

    View details for PubMedID 20435763

    View details for PubMedCentralID PMC2897429

  • Role of nitric oxide in Salmonella typhimurium-mediated cancer cell killing BMC CANCER Barak, Y., Schreiber, F., Thorne, S. H., Contag, C. H., deBeer, D., Matin, A. 2010; 10


    Bacterial targeting of tumours is an important anti-cancer strategy. We previously showed that strain SL7838 of Salmonella typhimurium targets and kills cancer cells. Whether NO generation by the bacteria has a role in SL7838 lethality to cancer cells is explored. This bacterium has the mechanism for generating NO, but also for decomposing it.Mechanism underlying Salmonella typhimurium tumour therapy was investigated through in vitro and in vivo studies. NO measurements were conducted either by chemical assays (in vitro) or using Biosensors (in vivo). Cancer cells cytotoxic assay were done by using MTS. Bacterial cell survival and tumour burden were determined using molecular imaging techniques.SL7838 generated nitric oxide (NO) in anaerobic cell suspensions, inside infected cancer cells in vitro and in implanted 4T1 tumours in live mice, the last, as measured using microsensors. Thus, under these conditions, the NO generating pathway is more active than the decomposition pathway. The latter was eliminated, in strain SL7842, by the deletion of hmp- and norV genes, making SL7842 more proficient at generating NO than SL7838. SL7842 killed cancer cells more effectively than SL7838 in vitro, and this was dependent on nitrate availability. This strain was also ca. 100% more effective in treating implanted 4T1 mouse tumours than SL7838.NO generation capability is important in the killing of cancer cells by Salmonella strains.

    View details for DOI 10.1186/1471-2407-10-146

    View details for Web of Science ID 000277802300001

    View details for PubMedID 20398414

    View details for PubMedCentralID PMC2868810

  • Visualizing Implanted Tumors in Mice with Magnetic Resonance Imaging Using Magnetotactic Bacteria CLINICAL CANCER RESEARCH Benoit, M. R., Mayer, D., Barak, Y., Chen, I. Y., Hu, W., Cheng, Z., Wang, S. X., Spielman, D. M., Gambhir, S. S., Matin, A. 2009; 15 (16): 5170-5177


    To determine if magnetotactic bacteria can target tumors in mice and provide positive contrast for visualization using magnetic resonance imaging.The ability of the magnetotactic bacterium, Magnetospirillum magneticum AMB-1 (referred to from here as AMB-1), to confer positive magnetic resonance imaging contrast was determined in vitro and in vivo. For the latter studies, AMB-1 were injected either i.t. or i.v. Bacterial growth conditions were manipulated to produce small (approximately 25-nm diameter) magnetite particles, which were observed using transmission electron microscopy. Tumor targeting was confirmed using 64Cu-labeled bacteria and positron emission tomography and by determination of viable cell counts recovered from different organs and the tumor.We show that AMB-1 bacteria with small magnetite particles generate T1-weighted positive contrast, enhancing in vivo visualization by magnetic resonance imaging. Following i.v. injection of 64Cu-labeled AMB-1, positron emission tomography imaging revealed increasing colonization of tumors and decreasing infection of organs after 4 hours. Viable cell counts showed that, by day 6, the bacteria had colonized tumors but were cleared completely from other organs. Magnetic resonance imaging showed a 1.22-fold (P = 0.003) increased positive contrast in tumors on day 2 and a 1.39-fold increase (P = 0.0007) on day 6.Magnetotactic bacteria can produce positive magnetic resonance imaging contrast and colonize mouse tumor xenografts, providing a potential tool for improved magnetic resonance imaging visualization in preclinical and translational studies to track cancer.

    View details for DOI 10.1158/1078-0432.CCR-08-3206

    View details for Web of Science ID 000269024900019

    View details for PubMedID 19671860

    View details for PubMedCentralID PMC3409839

  • CNOB/ChrR6, a new prodrug enzyme cancer chemotherapy MOLECULAR CANCER THERAPEUTICS Thorne, S. H., Barak, Y., Liang, W., Bachmann, M. H., Rao, J., Contag, C. H., Matin, A. 2009; 8 (2): 333-341


    We report the discovery of a new prodrug, 6-chloro-9-nitro-5-oxo-5H-benzo(a)phenoxazine (CNOB). This prodrug is efficiently activated by ChrR6, the highly active prodrug activating bacterial enzyme we have previously developed. The CNOB/ChrR6 therapy was effective in killing several cancer cell lines in vitro. It also efficiently treated tumors in mice with up to 40% complete remission. 9-Amino-6-chloro-5H-benzo(a)phenoxazine-5-one (MCHB) was the only product of CNOB reduction by ChrR6. MCHB binds DNA; at nonlethal concentration, it causes cell accumulation in the S phase, and at lethal dose, it induces cell surface Annexin V and caspase-3 and caspase-9 activities. Further, MCHB colocalizes with mitochondria and disrupts their electrochemical potential. Thus, killing by CNOB involves MCHB, which likely induces apoptosis through the mitochondrial pathway. An attractive feature of the CNOB/ChrR6 regimen is that its toxic product, MCHB, is fluorescent. This feature proved helpful in in vitro studies because simple fluorescence measurements provided information on the kinetics of CNOB activation within the cells, MCHB killing mechanism, its generally efficient bystander effect in cells and cell spheroids, and its biodistribution. The emission wavelength of MCHB also permitted its visualization in live animals, allowing noninvasive qualitative imaging of MCHB in mice and the tumor microenvironment. This feature may simplify exploration of barriers to the penetration of MCHB in tumors and their amelioration.

    View details for DOI 10.1158/1535-7163.MCT-08-0707

    View details for Web of Science ID 000263397300008

    View details for PubMedID 19190118

    View details for PubMedCentralID PMC2670992

  • Enzyme improvement in the absence of structural knowledge: a novel statistical approach ISME JOURNAL Barak, Y., Nov, Y., Ackerley, D. F., Matin, A. 2008; 2 (2): 171-179


    Most existing methods for improving protein activity are laborious and costly, as they either require knowledge of protein structure or involve expression and screening of a vast number of protein mutants. We describe here a successful first application of a novel approach, which requires no structural knowledge and is shown to significantly reduce the number of mutants that need to be screened. In the first phase of this study, around 7000 mutants were screened through standard directed evolution, yielding a 230-fold improvement in activity relative to the wild type. Using sequence analysis and site-directed mutagenesis, an additional single mutant was then produced, with 500-fold improved activity. In the second phase, a novel statistical method for protein improvement was used; building on data from the first phase, only 11 targeted additional mutants were produced through site-directed mutagenesis, and the best among them achieved a >1500-fold improvement in activity over the wild type. Thus, the statistical model underlying the experiment was validated, and its predictions were shown to reduce laboratory labor and resources.

    View details for DOI 10.1038/ismej.2007.100

    View details for Web of Science ID 000253492900005

    View details for PubMedID 18253133

  • Therapeutic implications of nitric oxide generation by tumor-targeting Salmonella typhimurium strains. Barak, Y., Schreiber, F., Thorne, S. H., Contag, C. H., de-Beer, D., Matin, A. C. AMER ASSOC CANCER RESEARCH. 2007: 3559S
  • Role of the rapA gene in controlling antibiotic resistance of Escherichia coli biofilms ANTIMICROBIAL AGENTS AND CHEMOTHERAPY Lynch, S. V., Dixon, L., Benoit, M. R., Brodie, E. L., Keyhan, M., Hu, P., Ackerley, D. F., Andersen, G. L., Matin, A. 2007; 51 (10): 3650-3658


    By using a high-throughput screening method, a mutant of a uropathogenic Escherichia coli strain affected in the rapA gene was isolated. The mutant formed normal-architecture biofilms but showed decreased penicillin G resistance, although the mutation did not affect planktonic cell resistance. Transcriptome analysis showed that 22 genes were down-regulated in the mutant biofilm. One of these genes was yhcQ, which encodes a putative multidrug resistance pump. Mutants with mutations in this gene also formed biofilms with decreased resistance, although the effect was less pronounced than that of the rapA mutation. Thus, an additional mechanism(s) controlled by a rapA-regulated gene(s) was involved in wild-type biofilm resistance. The search for this mechanism was guided by the fact that another down-regulated gene in rapA biofilms, yeeZ, is suspected to be involved in extra cell wall-related functions. A comparison of the biofilm matrix of the wild-type and rapA strains revealed decreased polysaccharide quantities and coverage in the mutant biofilms. Furthermore, the (fluorescent) functional penicillin G homologue Bocillin FL penetrated the mutant biofilms more readily. The results strongly suggest a dual mechanism for the wild-type biofilm penicillin G resistance, retarded penetration, and effective efflux. The results of studies with an E. coli K-12 strain pointed to the same conclusion. Since efflux and penetration can be general resistance mechanisms, tests were conducted with other antibiotics. The rapA biofilm was also more sensitive to norfloxacin, chloramphenicol, and gentamicin.

    View details for DOI 10.1128/AAC.00601-07

    View details for Web of Science ID 000249794800025

    View details for PubMedID 17664315

    View details for PubMedCentralID PMC2043260

  • Evolved high activity enzymes for enhancing combined bacterial chromate and uranyl bioremediation Matin, A. C., Barak, Y., Ackerley, D., Nov, Y., Francis, A. J., Dodge, C. AMER CHEMICAL SOC. 2007: 581
  • Escherichia coli biofilms formed under low-shear modeled microgravity in a ground-based system APPLIED AND ENVIRONMENTAL MICROBIOLOGY Lynch, S. V., Mukundakrishnan, K., Benoit, M. R., Ayyaswamy, P. S., Matin, A. 2006; 72 (12): 7701-7710


    Bacterial biofilms cause chronic diseases that are difficult to control. Since biofilm formation in space is well documented and planktonic cells become more resistant and virulent under modeled microgravity, it is important to determine the effect of this gravity condition on biofilms. Inclusion of glass microcarrier beads of appropriate dimensions and density with medium and inoculum, in vessels specially designed to permit ground-based investigations into aspects of low-shear modeled microgravity (LSMMG), facilitated these studies. Mathematical modeling of microcarrier behavior based on experimental conditions demonstrated that they satisfied the criteria for LSMMG conditions. Experimental observations confirmed that the microcarrier trajectory in the LSMMG vessel concurred with the predicted model. At 24 h, the LSMMG Escherichia coli biofilms were thicker than their normal-gravity counterparts and exhibited increased resistance to the general stressors salt and ethanol and to two antibiotics (penicillin and chloramphenicol). Biofilms of a mutant of E. coli, deficient in sigma(s), were impaired in developing LSMMG-conferred resistance to the general stressors but not to the antibiotics, indicating two separate pathways of LSMMG-conferred resistance.

    View details for DOI 10.1128/AEM.01294-06

    View details for Web of Science ID 000242681300035

    View details for PubMedID 17028231

    View details for PubMedCentralID PMC1694224

  • Analysis of novel soluble chromate and uranyl reductases and generation of an improved enzyme by directed evolution APPLIED AND ENVIRONMENTAL MICROBIOLOGY Barak, Y., Ackerley, D. F., Dodge, C. J., Banwari, L., Alex, C., Francis, A. J., Matin, A. 2006; 72 (11): 7074-7082


    Most polluted sites contain mixed waste. This is especially true of the U.S. Department of Energy (DOE) waste sites which hold a complex mixture of heavy metals, radionuclides, and organic solvents. In such environments enzymes that can remediate multiple pollutants are advantageous. We report here evolution of an enzyme, ChrR6 (formerly referred to as Y6), which shows a markedly enhanced capacity for remediating two of the most serious and prevalent DOE contaminants, chromate and uranyl. ChrR6 is a soluble enzyme and reduces chromate and uranyl intracellularly. Thus, the reduced product is at least partially sequestered and nucleated, minimizing the chances of reoxidation. Only one amino acid change, (Tyr)128(Asn), was responsible for the observed improvement. We show here that ChrR6 makes Pseudomonas putida and Escherichia coli more efficient agents for bioremediation if the cellular permeability barrier to the metals is decreased.

    View details for DOI 10.1128/AEM.01334-06

    View details for Web of Science ID 000242003800027

    View details for PubMedID 17088379

    View details for PubMedCentralID PMC1636143

  • Effect of chromate stress on Escherichia coli K-12 JOURNAL OF BACTERIOLOGY Ackerley, D. F., Barak, Y., Lynch, S. V., Curtin, J., Matin, A. 2006; 188 (9): 3371-3381


    The nature of the stress experienced by Escherichia coli K-12 exposed to chromate, and mechanisms that may enable cells to withstand this stress, were examined. Cells that had been preadapted by overnight growth in the presence of chromate were less stressed than nonadapted controls. Within 3 h of chromate exposure, the latter ceased growth and exhibited extreme filamentous morphology; by 5 h there was partial recovery with restoration of relatively normal cell morphology. In contrast, preadapted cells were less drastically affected in their morphology and growth. Cellular oxidative stress, as monitored by use of an H2O2-responsive fluorescent dye, was most severe in the nonadapted cells at 3 h postinoculation, lower in the partially recovered cells at 5 h postinoculation, and lower still in the preadapted cells. Chromate exposure depleted cellular levels of reduced glutathione and other free thiols to a greater extent in nonadapted than preadapted cells. In both cell types, the SOS response was activated, and levels of proteins such as SodB and CysK, which can counter oxidative stress, were increased. Some mutants missing antioxidant proteins (SodB, CysK, YieF, or KatE) were more sensitive to chromate. Thus, oxidative stress plays a major role in chromate toxicity in vivo, and cellular defense against this toxicity involves activation of antioxidant mechanisms. As bacterial chromate bioremediation is limited by the toxicity of chromate, minimizing oxidative stress during bacterial chromate reduction and bolstering the capacity of these organisms to deal with this stress will improve their effectiveness in chromate bioremediation.

    View details for DOI 10.1128/JB.188.9.3371-3381.2006

    View details for Web of Science ID 000237171200026

    View details for PubMedID 16621832

  • New enzyme for reductive cancer chemotherapy, YieF, and its improvement by directed evolution MOLECULAR CANCER THERAPEUTICS Barak, Y., Thorne, S. H., Ackerley, D. F., Lynch, S. V., Contag, C. H., Matin, A. 2006; 5 (1): 97-103


    Reductive prodrugs, mitomycin C and 5-aziridinyl-2,4-dinitrobenzamide (CB 1954), are nontoxic in their native form but become highly toxic upon reduction. Their effectiveness in cancer chemotherapy can be enhanced by delivering to tumors enzymes with improved prodrug reduction kinetics. We report the discovery of a new prodrug-reducing enzyme, YieF, from Escherichia coli, and the improvement of its kinetics for reducing mitomycin C and CB 1954. A YieF-derived enzyme, Y6, killed HeLa spinner cells with >or=5-fold efficiency than the wild-type enzymes, YieF and NfsA, at a variety of drug and enzyme concentrations and incubation times. With adhered HeLa cells and Salmonella typhimurium SL 7838 bacteria as enzyme delivery vehicle, at least an order of magnitude less of Y6-producing bacteria were required to kill >90% of tumor cells compared with bacteria expressing the wild-type enzymes, which at a comparable level killed < 5% of the cells. Thus, Y6 is a promising enzyme for use in cancer chemotherapy, and Salmonella strain SL 7838, which specifically targets tumors, may be used to deliver the prodrug-activating enzymes to tumors.

    View details for DOI 10.1158/1535-7163.MCT-05-0365

    View details for Web of Science ID 000234772900011

    View details for PubMedID 16432167

  • ChrR, a soluble quinone reductase of Pseudomonas putida that defends against H2O2 JOURNAL OF BIOLOGICAL CHEMISTRY Gonzalez, C. F., Ackerley, D. F., Lynch, S. V., Matin, A. 2005; 280 (24): 22590-22595


    Most bacteria contain soluble quinone-reducing flavoenzymes. However, no biological benefit for this activity has previously been demonstrated. ChrR of Pseudomonas putida is one such enzyme that has also been characterized as a chromate reductase; yet we propose that it is the quinone-reducing activity of ChrR that has the greatest biological significance. ChrR reduces quinones by simultaneous two-electron transfer, avoiding formation of highly reactive semiquinone intermediates and producing quinols that promote tolerance of H(2)O(2). Expression of chrR was induced by H(2)O(2), and levels of chrR expression in overexpressing, wild type, and knock-out mutant strains correlated with the H(2)O(2) tolerance and scavenging ability of each strain. The chrR expression level also correlated with intracellular H(2)O(2) levels as measured by protein carbonylation assays and fluorescence-activated cell scanning analysis with the H(2)O(2)-responsive dye H(2)DCFDA. Thus, enhancing the activity of ChrR in a chromate-remediating bacterial strain may not only increase the rate of chromate transformation, it may also augment the capacity of these cells to withstand the unavoidable production of H(2)O(2) that accompanies chromate reduction.

    View details for DOI 10.1074/jbc.M501654200

    View details for Web of Science ID 000229741800007

    View details for PubMedID 15840577

  • Investigating the threat of bacteria grown in space ASM NEWS Matin, A., Lynch, S. V. 2005; 71 (5): 235-240
  • Role and regulation of sigma(s) in general resistance conferred by low-shear simulated microgravity in Escherichia coli JOURNAL OF BACTERIOLOGY Lynch, S. V., Brodie, E. L., Matin, A. 2004; 186 (24): 8207-8212


    Life on Earth evolved in the presence of gravity, and thus it is of interest from the perspective of space exploration to determine if diminished gravity affects biological processes. Cultivation of Escherichia coli under low-shear simulated microgravity (SMG) conditions resulted in enhanced stress resistance in both exponential- and stationary-phase cells, making the latter superresistant. Given that microgravity of space and SMG also compromise human immune response, this phenomenon constitutes a potential threat to astronauts. As low-shear environments are encountered by pathogens on Earth as well, SMG-conferred resistance is also relevant to controlling infectious disease on this planet. The SMG effect resembles the general stress response on Earth, which makes bacteria resistant to multiple stresses; this response is sigma s dependent, irrespective of the growth phase. However, SMG-induced increased resistance was dependent on sigma s only in stationary phase, being independent of this sigma factor in exponential phase. sigma s concentration was some 30% lower in exponential-phase SMG cells than in normal gravity cells but was twofold higher in stationary-phase SMG cells. While SMG affected sigma s synthesis at all levels of control, the main reasons for the differential effect of this gravity condition on sigma s levels were that it rendered the sigma protein less stable in exponential phase and increased rpoS mRNA translational efficiency. Since sigma s regulatory processes are influenced by mRNA and protein-folding patterns, the data suggest that SMG may affect these configurations.

    View details for DOI 10.1128/JB.186.24.8207-8212.2004

    View details for Web of Science ID 000225670300006

    View details for PubMedID 15576768

  • Mechanism of chromate reduction by the Escherichia coli protein, NfsA, and the role of different chromate reductases in minimizing oxidative stress during chromate reduction ENVIRONMENTAL MICROBIOLOGY Ackerley, D. F., Gonzalez, C. F., Keyhan, M., Blake, R., Matin, A. 2004; 6 (8): 851-860


    Chromate [Cr(VI)] is a serious environmental pollutant, which is amenable to bacterial bioremediation. NfsA, the major oxygen-insensitive nitroreductase of Escherichia coli, is a flavoprotein that is able to reduce chromate to less soluble and less toxic Cr(III). We show that this process involves single-electron transfer, giving rise to a flavin semiquinone form of NfsA and Cr(V) as intermediates, which redox cycle, generating more reactive oxygen species (ROS) than a divalent chromate reducer, YieF. However, NfsA generates less ROS than a known one-electron chromate reducer, lipoyl dehydrogenase (LpDH), suggesting that NfsA employs a mixture of uni- and di-valent electron transfer steps. The presence of YieF, ChrR (another chromate reductase we previously characterized), or NfsA in an LpDH-catalysed chromate reduction reaction decreased ROS generation by c. 65, 40, or 20%, respectively, suggesting that these enzymes can pre-empt ROS generation by LpDH. We previously showed that ChrR protects Pseudomonas putida against chromate toxicity; here we show that NfsA or YieF overproduction can also increase the tolerance of E. coli to this compound.

    View details for DOI 10.1111/j.1462-2920.2004.00639.x

    View details for Web of Science ID 000222578600010

    View details for PubMedID 15250887

  • Chromate-reducing properties of soluble Flavoproteins from Pseudomonas putida and Escherichia coli APPLIED AND ENVIRONMENTAL MICROBIOLOGY Ackerley, D. F., Gonzalez, C. F., Park, C. H., Blake, R., Keyhan, A., Matin, A. 2004; 70 (2): 873-882


    Cr(VI) (chromate) is a toxic, soluble environmental contaminant. Bacteria can reduce chromate to the insoluble and less toxic Cr(III), and thus chromate bioremediation is of interest. Genetic and protein engineering of suitable enzymes can improve bacterial bioremediation. Many bacterial enzymes catalyze one-electron reduction of chromate, generating Cr(V), which redox cycles, generating excessive reactive oxygen species (ROS). Such enzymes are not appropriate for bioremediation, as they harm the bacteria and their primary end product is not Cr(III). In this work, the chromate reductase activities of two electrophoretically pure soluble bacterial flavoproteins--ChrR (from Pseudomonas putida) and YieF (from Escherichia coli)-were examined. Both are dimers and reduce chromate efficiently to Cr(III) (kcat/Km = approximately 2 x 10(4) M(-1) x s(-1)). The ChrR dimer generated a flavin semiquinone during chromate reduction and transferred >25% of the NADH electrons to ROS. However, the semiquinone was formed transiently and ROS diminished with time. Thus, ChrR probably generates Cr(V), but only transiently. Studies with mutants showed that ChrR protects against chromate toxicity; this is possibly because it preempts chromate reduction by the cellular one-electron reducers, thereby minimizing ROS generation. ChrR is thus a suitable enzyme for further studies. During chromate reduction by YieF, no flavin semiquinone was generated and only 25% of the NADH electrons were transferred to ROS. The YieF dimer may therefore be an obligatory four-electron chromate reducer which in one step transfers three electrons to chromate and one to molecular oxygen. As a mutant lacking this enzyme could not be obtained, the role of YieF in chromate protection could not be directly explored. The results nevertheless suggest that YieF may be an even more suitable candidate for further studies than ChrR.

    View details for DOI 10.1128/AEM.70.2.873-882.2004

    View details for Web of Science ID 000188854900031

    View details for PubMedID 14766567

    View details for PubMedCentralID PMC348923

  • The stress response of Escherichia coli to conditions of simulated microgravity Abstracts of the 102nd General meeting of the American Society for Microbiology, Washington D.C., A. Matin., Lynch, S. V. 2003
  • Tetracycline rapidly reaches all the constituent cells of uropathogenic Escherichia coli biofilms ANTIMICROBIAL AGENTS AND CHEMOTHERAPY Stone, G., Wood, P., Dixon, L., Keyhan, M., Matin, A. 2002; 46 (8): 2458-2461


    We have developed a method for visualizing Escherichia coli cells that are exposed to tetracycline in a biofilm, based on a previous report that liposomes containing the E. coli TetR(B) protein fluoresce when exposed to this antibiotic. By our method, cells devoid of TetR(B) also exhibited tetracycline-dependent fluorescence. At 50 microg of tetracycline ml(-1), planktonic cells of a uropathogenic E. coli (UPEC) strain developed maximal fluorescence after 7.5 to 10 min of exposure. A similar behavior was exhibited by cells in a 24- or 48-h UPEC biofilm, as examined by confocal laser microscopy, regardless of whether they lined empty spaces or occupied densely packed regions. Further, a comparison of phase-contrast and fluorescent images of corresponding biofilm zones showed that all the cells fluoresced. Thus, all the biofilm cells were exposed to tetracycline and there were no pockets within the biofilm where the antibiotic failed to reach. It also appeared unlikely that niches of reduced exposure to the antibiotic existed within the biofilms.

    View details for DOI 10.1128/AAC.46.8.2458.2461.2002

    View details for Web of Science ID 000176968700021

    View details for PubMedID 12121918

  • A soluble flavoprotein contributes to chromate reduction and tolerance by Pseudomonas putida. Acta Biotechnology Ackerley DF, Park CH, Gonzalez CF, Keyhan M, Matin A 2002; 23: 233
  • The EmrR protein represses the Escherichia coli emrRAB multidrug resistance operon by directly binding to its promoter region ANTIMICROBIAL AGENTS AND CHEMOTHERAPY Xiong, A., Gottman, A., PARK, C., Baetens, M., Pandza, S., Matin, A. 2000; 44 (10): 2905-2907


    EmrR negatively regulates the transcription of the multidrug resistance pump-encoding operon, emrRAB, by binding to its regulatory region. The binding site spans the promoter and the downstream sequence up to the transcriptional start site of the operon. Structurally unrelated drugs that induce the pump interfere with this binding.

    View details for Web of Science ID 000089402300059

    View details for PubMedID 10991887

  • Purification to homogeneity and characterization of a novel Pseudomonas putida chromate reductase APPLIED AND ENVIRONMENTAL MICROBIOLOGY Park, C. H., Keyhan, M., Wielinga, B., Fendorf, S., Matin, A. 2000; 66 (5): 1788-1795


    Cr(VI) (chromate) is a widespread environmental contaminant. Bacterial chromate reductases can convert soluble and toxic chromate to the insoluble and less toxic Cr(III). Bioremediation can therefore be effective in removing chromate from the environment, especially if the bacterial propensity for such removal is enhanced by genetic and biochemical engineering. To clone the chromate reductase-encoding gene, we purified to homogeneity (>600-fold purification) and characterized a novel soluble chromate reductase from Pseudomonas putida, using ammonium sulfate precipitation (55 to 70%), anion-exchange chromatography (DEAE Sepharose CL-6B), chromatofocusing (Polybuffer exchanger 94), and gel filtration (Superose 12 HR 10/30). The enzyme activity was dependent on NADH or NADPH; the temperature and pH optima for chromate reduction were 80 degrees C and 5, respectively; and the K(m) was 374 microM, with a V(max) of 1.72 micromol/min/mg of protein. Sulfate inhibited the enzyme activity noncompetitively. The reductase activity remained virtually unaltered after 30 min of exposure to 50 degrees C; even exposure to higher temperatures did not immediately inactivate the enzyme. X-ray absorption near-edge-structure spectra showed quantitative conversion of chromate to Cr(III) during the enzyme reaction.

    View details for Web of Science ID 000086805500003

    View details for PubMedID 10788340

  • The G-protein FlhF has a role in polar flagellar placement and general stress response induction in Pseudomonas putida MOLECULAR MICROBIOLOGY Pandza, S., Baetens, M., Park, C. H., Au, T., Keyhan, M., Matin, A. 2000; 36 (2): 414-423


    The flhF gene of Pseudomonas putida, which encodes a GTP-binding protein, is part of the flagellar-motility-chemotaxis operon. Its disruption leads to a random flagellar arrangement in the mutant (MK107) and loss of directional motility in contrast to the wild type, which has polar flagella. The return of a normal flhF allele restores polar flagella and normal motility to MK107; its overexpression triples the flagellar number but does not restore directional motility. As FlhF is homologous to the receptor protein of the signal recognition particle (SRP) pathway of membrane protein translocation, this pathway may have a role in polar flagellar placement in P. putida. MK107 is also compromised in the development of the starvation-induced general stress resistance (SGSR) and effective synthesis of several starvation and exponential phase proteins. While somewhat increased protein secretion in MK107 may contribute to its SGSR impairment, the altered protein synthesis pattern also appears to have a role.

    View details for Web of Science ID 000086980800014

    View details for PubMedID 10792727

  • How can archaea cope with extreme acidity? BACTERIAL RESPONSE TO PH Schafer, G., Krulwich, T. A., Poole, R. K., Padan, E., Konings, W. N., Skulachev, V., Fillingame, R. H., Matin, A., Dimroth, P., Booth, I. R., Bogachev, A., Cook, G. M., Dilworth, M. J., Epstein, W. 1999; 221: 131-151


    Archaea are the most extremophilic of the acidophilic microbes, combining, in many cases, acidophilicity with hyperthermophilicity. They form one of the three branches of the phylogenetic tree, and they are specifically found within the so-called crenarchaeota, typical members of which thrive at pH 1-3 and at temperatures of 75 degrees C to nearly 100 degrees C. Despite this, these cells can maintain a near neutral cytosol, and they use H+ for chemiosmotic coupling of ADP phosphorylation. These phenomena require efficient exclusion and disposal of protons. This is achieved by multiple synergistic mechanisms that act in parallel. One strategy is to use bipolar tetraether lipids as a matrix of their plasma membranes, providing low ion permeabilities, even at high temperatures. Additionally, an inverted membrane potential can help to balance a large pH gradient of up to 4 at a proton motive force of delta p = 140-180 mV. This is not a general rule, because in several species the membrane potential contributes only minimally. Also, local buffering capacity and charge profiles across the membrane may significantly influence adaptation to bulk phase acidity. Neither complex I nor complex III electron transport-coupled proton pump equivalents have been found in aerobic archaea. Only terminal oxidases seem to provide either H+ pumping or the generation of a proton gradient by chemical charge separation. Organization, redox centres and primary structures of some archaeal terminal quinol oxidase complexes are known and will be discussed. Much less is known about anaerobic sulfur reducers. For those a possible mechanism for proton exclusion is proposed.

    View details for Web of Science ID 000081620400009

    View details for PubMedID 10207917

  • Acid tolerance induced by metabolites and secreted proteins, and how tolerance can be counteracted Symposium on Bacterial Responses to pH Rowbury, R. J., Foster, J. W., Konings, W. N., Matin, A., Poole, R. K., Schafer, G., Glenn, A. R., Booth, I. R., Park, S. F., Slonczewski, J. L., Epstein, W., Cook, G. M. JOHN WILEY & SONS LTD. 1999: 93–111


    Several metabolites and salts including glucose, L-glutamate, L-aspartate, FeCl3, KCl and L-proline induce acid tolerance at neutral external pH (pHo) in log phase Escherichia coli. For induction by glucose and L-glutamate, the processes are independent of integration host factor (IHF), H-NS, CysB, ferric uptake regulator (Fur) and RelA. For most of the above, tolerance does not appear if induction occurs and NaCl, sucrose, SDS or DOC are present. For several responses, cAMP inhibits induction. For many established acid tolerance and sensitization processes, including those tolerance responses switched on at pH 5.0 and by glucose, glutamate or aspartate, induction is associated with secretion of extracellular induction proteins. These proteins bring about the response if added to organisms under normally non-inducing conditions. Secreted components also influence inherent acid tolerances and sensitivities. Analysis of some established tolerance responses indicates that induction is a two-stage process, secreted extracellular proteins playing an obligate role in induction. For example, the functioning of the acid-induced medium protein(s) is essential for acid habituation at pHo 5.0. It seems likely that such two-stage mechanisms are essential for many inducible processes in bacteria.

    View details for Web of Science ID 000081620400007

    View details for PubMedID 10207915

  • Acid and base regulation in the proteome of Escherichia coli Symposium on Bacterial Responses to pH Slonczewski, J. L., Blankenhorn, D., Foster, J. W., Matin, A., Booth, I. R., Stock, J. B., Skulachev, V., Rowbury, R. J., Konings, W. N., Bennett, G. N., Kobayashi, H., Fillingame, R. H., Schafer, G., Poole, R. K., Krulwich, T. A., Cook, G. M., Quivey, R. G. JOHN WILEY & SONS LTD. 1999: 75–92


    Acid and base conditions have many significant effects on the growth of Escherichia coli. External and internal pH perturbations induce different classes of genes. pH-dependent regulation of genes intersects with other regulatory responses, e.g. oxygen level or osmolarity. 2D electrophoretic gels were used to compare global patterns of protein induction in Escherichia coli grown in complex media buffered at the acid or alkaline ends of the pH range for growth (pH 4.4 vs. pH 9.1). Preliminary results indicate new classes of acid- and base-dependent regulation, in some cases highly dependent on oxygen level. Other proteins are induced strongly at both extremes of pH, compared to pH 7. Current work continues to dissect the relationship between effects of pH, oxygen level and osmolarity.

    View details for Web of Science ID 000081620400006

    View details for PubMedID 10207914

  • pH sensing in bacterial chemotaxis Symposium on Bacterial Responses to pH Levit, M. N., Stock, J. B., Foster, J. W., Matin, A., Fillingame, R. H., Konings, W. N., Slonczewski, J. L., Padan, E., Booth, I. R. JOHN WILEY & SONS LTD. 1999: 38–54


    Bacteria are able to sense a broad range of chemical and energetic stimuli and modulate their swimming behaviour to migrate to more favourable environments. Signal transduction in bacterial chemotaxis is mediated by a two-component system composed of a protein histidine kinase, CheA, and a response regulator, CheY. The phosphorylated response regulator, P approximately CheY, binds to a protein at the flagellar motor, FliM, to cause reversals in flagellar motor rotation. The level of P approximately CheY is controlled by the activity of the kinase CheA, which is in turn regulated by membrane receptors at the cell surface. Membrane receptors such as the aspartate receptor, Tar, are composed of two distinct regions: an extracellular sensing domain that binds stimulatory ligands, aspartate in the case of Tar; and an intracellular signalling domain that forms a complex with the protein kinase CheA. What is the mechanism of transmembrane signalling? How does aspartate binding to the sensing domain at the outside surface of the membrane translate into a change in kinase activity at the membrane cytosol interface? Recent results suggest that the mechanism depends on perturbations in lateral packing within an extensive array of receptors localized to patches at the cell poles. Receptor patching appears to depend on higher-order associations with the kinase CheA as well as an adaptor protein, CheW. It is difficult to assess the locus of pH effects within the context of even a simple signal transduction system like that involved in bacterial chemotaxis. Previous results with mutant strains have indicated that the serine receptor, Tsr, is critical for pH sensing, but in vitro results do not support such a straightforward interpretation of the genetic data.

    View details for Web of Science ID 000081620400004

    View details for PubMedID 10207912

  • pH tolerance in Bacillus: alkaliphiles versus non-alkaliphiles Symposium on Bacterial Responses to pH Krulwich, T. A., Guffanti, A. A., Ito, M., Quivey, R. G., Skulachev, V., Matin, A., Stock, J. B., Konings, W. N., Glenn, A. R. JOHN WILEY & SONS LTD. 1999: 167–182


    Monovalent cation/proton antiporters that catalyse electrogenic uptake of H+ in exchange for cytoplasmic K+ and/or Na+ are centrally involved in bacterial pH homeostasis under alkaline challenge. Systematic attempts have identified some, but not yet all, of the genes encoding such antiporters that participate in pH homeostasis in the neutrophilic Bacillus subtilis and the extremely alkaliphilic Bacillus firmus OF4. In each organism there are at least three distinct antiporters involved in pH homeostasis. They differ in cation requirement, with pH homeostasis specifically utilizing Na+/H+ antiport in the alkaliphile and using either Na+ or K+/H+ antiport in B. subtilis. Some of the antiporters involved in pH homeostasis are constitutive and are in place to respond to sudden pH shifts, but there is also an inducible component. At least two sets of homologous antiporters (NhaC and Mrp/Pha) function in both alkaliphiles and neutrophiles. An additional antiporter of a different transport protein family, the Gram-positive tetracycline-metal/H+ antiporter, is important in pH homeostasis in B. subtilis but has not yet been shown to be present in any alkaliphile. There are also differences outside of the antiporters themselves that contribute to the greater capacity of the alkaliphiles for pH homeostasis, including cation re-entry capacity and possible surface properties.

    View details for Web of Science ID 000081620400011

    View details for PubMedID 10207919

  • Final general discussion Symposium on Bacterial Responses to pH Bennett, G. N., Konings, W. N., Booth, I. R., Cook, G. M., Krulwich, T. A., Skulachev, V., Schafer, G., Epstein, W., Stock, J. B., Poole, R. K., Slonczewski, J. L., Glenn, A. R., Dilworth, M. J., Padan, E., Matin, A., Moir, A., Quivey, R. G. JOHN WILEY & SONS LTD. 1999: 246–250
  • The Escherichia coli starvation gene cstC is involved in amino acid catabolism JOURNAL OF BACTERIOLOGY Fraley, C. D., Kim, J. H., McCann, M. P., Matin, A. 1998; 180 (16): 4287-4290


    Escherichia coli strains mutant in the starvation gene cstC grow normally in a mineral salts medium but are impaired in utilizing amino acids as nitrogen sources. They are also compromised in starvation survival, where amino acid catabolism is important. The cstC gene encodes a 406-amino-acid protein that closely resembles the E. coli ArgD protein, which is involved in arginine biosynthesis. We postulate that CstC is a counterpart of ArgD in an amino acid catabolic pathway. The cstC upstream region contains several regulatory consensus sequences. Both sigmaS and sigma54 promoters are probably involved in cstC transcription and appear to compete with each other, presumably to match cstC expression to the cellular amino acid catabolic needs.

    View details for Web of Science ID 000075328400036

    View details for PubMedID 9696780

  • H-NS protein represses transcription of the lux systems of Vibrio fischeri and other luminous bacteria cloned into Escherichia coli CURRENT MICROBIOLOGY Ulitzur, S., Matin, A., Fraley, C., Meighen, E. 1997; 35 (6): 336-342


    High expression in Escherichia coli of the lux system cistron of a luminous bacteria under its own control has been accomplished only for the Vibrio fischeri lux system at high cell density. Mutation of the hns gene in E. coli has resulted in strong expression of the V. fischeri lux system at low cell density even in an rpoS-deleted strain of E. coli that emits very low levels of luminescence. The E. coli double mutant, MC4110 hns::kan rpoS::tet carrying the lux system of V. fischeri, developed high luminescence from the very early stages of cellular growth, regardless of the presence of deletion mutations in the luxI or luxR genes. Moreover, autoinducer synthesis was restored in the double mutant with the luxR-deleted system. plac-controlled V. fischeri luxCDABE genes missing luxI and luxR were dim in E. coli rpoS mutant cells, but had wild-type levels of light in the hns-deleted strain [MC4110 hns rpoS], showing that expression was independent of lux regulators in the absence of H-NS. DNA gyrase inhibitors and DNA intercalating agents also brought about the restoration of luminescence in the rpoS-deficient strain. High expression of the lux systems of Vibrio harveyi, Photobacterium leiognathi, and Xenorhabdus luminescens in E. coli MC4110 hns rpoS cells compared with that in wild-type or rpoS mutants was also accomplished. Taken together, these data suggest that the H-NS protein inhibits transcription in E. coli of the lux systems of all or most luminous bacteria at the luxC gene as well as in the luxRI region of the V. fischeri lux operon. These DNA regions are highly enriched with homopolymeric stretches of poly d(A) and poly d(T) characterizing curved DNA, a preferable site of H-NS binding. The significance of the new findings in understanding the regulatory control of the bacterial lux system is discussed.

    View details for Web of Science ID A1997YG68200004

    View details for PubMedID 9353217

  • The sigma(s) level in starving Escherichia coli cells increases solely as a result of its increased stability, despite decreased synthesis MOLECULAR MICROBIOLOGY Zgurskaya, H. I., Keyhan, M., Matin, A. 1997; 24 (3): 643-651


    The sigma S level in starving (stationary phase) Escherichia coli cells increases four-to sixfold following growth in a defined or a complex medium. Chemostat-grown cells, subjected to increasing carbon starvation, also become progressively richer in sigma S content. These increases occur despite reduced transcription of the sigma S-encoding gene, rpoS, and translation of rpoS mRNA, and result solely from a large increase in the stability of the sigma protein. Previous results, based on rpoS::lacZ transcriptional and translational fusions, and on methionine incorporation in sigma S, had suggested increased synthesis of sigma S in starving cells. Alternative explanations for these results consistent with the conclusions of this paper are discussed.

    View details for Web of Science ID A1997XA65100018

    View details for PubMedID 9179856

  • A bacterial model system for understanding multi-drug resistance MICROBIAL DRUG RESISTANCE Saier, M. H., Paulsen, I. T., Matin, A. 1997; 3 (4): 289-295


    Mankind stands at the crossroads, recognizing the need for a radical change in bacterial disease management. The development of several antimicrobial agents in the 1940s and 1950s allowed man to gain the upper hand in controlling these diseases. However, the horizon is now clouded by the activation in bacteria of cryptic multi-drug resistance (MDR) genes and the spread of plasmid- and integron-born MDR genes through bacterial populations. Unless remedial measures are taken, nearly all currently available antimicrobial agents are likely to soon lose their efficacies. We briefly review the bacterial MDR phenomenon and focus on a recently emerging family of small multi-drug resistance (SMR) pumps which may provide an ideal model system for understanding the MDR phenomenon in general.

    View details for Web of Science ID 000071196200001

    View details for PubMedID 9442481

  • Role of alternate sigma factors in starvation protein synthesis - Novel mechanisms of catabolite repression RESEARCH IN MICROBIOLOGY Matin, A. 1996; 147 (6-7): 494-505

    View details for Web of Science ID A1996VN56100009

    View details for PubMedID 9084761

  • Differential regulation of the mcb and emr operons of Escherichia coli: Role of mcb in multidrug resistance ANTIMICROBIAL AGENTS AND CHEMOTHERAPY Lomovskaya, O., Kawai, F., Matin, A. 1996; 40 (4): 1050-1052


    The mcb operon (which is responsible for microcin B17 production) and the emr operon (which encodes a multidrug resistance pump) share a common negative regulator, EmrR. Nevertheless, compounds that induce the emr operon repress the mcb operon. The pump dedicated to microcin B17 extrusion can also protect the calls against sparfloxacin and other toxic compounds.

    View details for Web of Science ID A1996UD24800048

    View details for PubMedID 8849229

  • Capacity of Helicobacter pylori to generate ionic gradients at low pH is similar to that of bacteria which grow under strongly acidic conditions INFECTION AND IMMUNITY Matin, A., Zychlinsky, E., Keyhan, M., Sachs, G. 1996; 64 (4): 1434-1436


    Helicobacter pylori colonized the highly acidic human gastric mucosa. At pH 3.0 to 7.0, this bacterium maintained a nearly neutral internal pH. Its membrane potential changed reciprocally with the pH gradient so that a relatively constant proton motive force was maintained. Possible, the capacity to maintain an appropriate transmembrane ionic gradient at a low pH contributes to the pathogenic propensities of this bacterium.

    View details for Web of Science ID A1996UC31400051

    View details for PubMedID 8606113

  • Regulation of Escherichia coli starvation sigma factor (sigma(s)) by ClpXP protease JOURNAL OF BACTERIOLOGY Schweder, T., Lee, K. H., Lomovskaya, O., Matin, A. 1996; 178 (2): 470-476


    In Escherichia coli, starvation (stationary-phase)-mediated differentiation involves 50 or more genes and is triggered by an increase in cellular sigma s levels. Western immunoblot analysis showed that in mutants lacking the protease ClpP or its cognate ATPase-containing subunit ClpX, sigma s levels of exponential-phase cells increased to those of stationary-phase wild-type cells. Lack of other potential partners of ClpP, i.e., ClpA or ClpB, or of Lon protease had no effect. In ClpXP-proficient cells, the stability of sigma s increased markedly in stationary-phase compared with exponential-phase cells, but in ClpP-deficient cells, sigma s became virtually completely stable in both phases. There was no decrease in ClpXP levels in stationary-phase wild-type cells. Thus, sigma s probably becomes more resistant to this protease in stationary phase. The reported sigma s-stabilizing effect of the hns mutation also was not due to decreased protease levels. Studies with translational fusions containing different lengths of sigma s coding region suggest that amino acid residues 173 to 188 of this sigma factor may directly or indirectly serve as at least part of the target for ClpXP protease.

    View details for Web of Science ID A1996TP52200020

    View details for PubMedID 8550468



    Volume 61, no. 9, p. 3323: the title of the article should read as shown above. [This corrects the article on p. 3323 in vol. 61.].

    View details for Web of Science ID A1995TC81100065

    View details for PubMedID 16535172



    The expression of much useful bacterial activity is facilitated by rapid growth. This coupling can create problems in bacterial fermentations and in situ bioremediation. In the latter process, for example, it necessitates addition of large amounts of nutrients to contaminated environments, such as aquifers. This approach, termed biostimulation, can be technically difficult. Moreover, the resulting in situ bacterial biomass production can have undesirable consequences. In an attempt to minimize coupling between expression of biodegradative activity and growth, we used Escherichia coli starvation promoters to control toluene monooxygenase synthesis. This enzyme complex can degrade the environmental contaminants trichloroethylene (TCE) and phenol. Totally starving cell suspensions of such strains degraded phenol and TCE. Furthermore, rapid conversions occurred in the postexponential batch or very slow growth (dilution) rate chemostat cultures, and the nutrient demand and biomass formation for transforming a given amount of TCE or phenol were reduced by 60 to 90%. Strong starvation promoters have recently been clones and characterized in environmentally relevant bacteria like Pseudomonas species; thus, starvation promoter-driven degradative systems can now be constructed in such bacteria and tested for in situ efficacy.

    View details for Web of Science ID A1995RT79800022

    View details for PubMedID 7574643



    The emrAB locus of Escherichia coli encodes a multidrug resistance pump that protects the cell from several chemically unrelated antimicrobial agents, e.g., the protonophores carbonyl cyanide m-chlorophenylhydrazone (CCCP) and tetrachlorosalicyl anilide and the antibiotics nalidixic acid and thiolactomycin. The mprA gene is located immediately upstream of this locus and was shown to be a repressor of microcin biosynthesis (I. del Castillo, J. M. Gomez, and F. Moreno, J. Bacteriol. 173:3924-3929, 1991). There is a putative transcriptional terminator sequence between the mprA and emrA genes. To locate the emr promoter, single-copy lacZ operon fusions containing different regions of the emr locus were made. Only fusions containing the mprA promoter region were expressed. mprA is thus the first gene of the operon, and we propose that it be renamed emrR. Overproduction of the EmrR protein (with a multicopy vector containing the cloned emrR gene) suppressed transcription of the emr locus. A mutation in the emrR gene led to overexpression of the EmrAB pump and increased resistance to antimicrobial agents. CCCP, nalidixic acid, and a number of other structurally unrelated chemicals induced expression of the emr genes, and the induction required EmrR. We conclude that emrRAB genes constitute an operon and that EmrR serves as a negative regulator of this operon. Some of the chemicals that induce the pump serve as its substrates, suggesting that their extrusion is the natural function of the pump.

    View details for Web of Science ID A1995QW52900013

    View details for PubMedID 7730261



    By using mini-Tn5 transposon mutagenesis, two mutants of Pseudomonas putida ATCC 12633 were isolated which showed a marked increase in their sensitivity to carbon starvation; these mutants are presumably affected in the Pex type of proteins that P. putida induces upon carbon starvation (M. Givskov, L. Eberl, and S. Molin, J. Bacteriol. 176:4816-4824, 1994). The affected genes in our mutants were induced about threefold upon carbon starvation. The promoter region of the starvation gene in the mutant MK107 possessed a strong sigma 54-type-promoter sequence, and deletion analysis suggested that this was the major promoter regulating expression; this was confirmed by transcript mapping in rpoN+ and rpoN mutant backgrounds. The deletion analysis implicated a sequence upstream of the sigma 54 promoter, as well as a region downstream of the transcription start site, in the functioning of the promoter. Two sigma 70-type Pribnow boxes were also detected in the promoter region, but their transcriptional activity in the wild type was very weak. However, in a sigma 54-deficient background, these promoters became stronger. The mechanism and possible physiological role of this phenomenon and the possibility that the sequence upstream of the sigma 54 promoter may have a role in carbon sensing are discussed.

    View details for Web of Science ID A1995QP81000028

    View details for PubMedID 7896711



    A reverse genetics approach was used to clone a pex starvation gene that codes for an 18-kDa polypeptide, designated PexB. Single-copy pexB-lacZ operon fusions were constructed to study transcriptional regulation and the promoter region of this gene. The induction by carbon starvation or osmotic stress was transcriptional and controlled by sigma 38 but was independent of this sigma factor by the oxidative stress; presumably, it was sigma 70 mediated under the latter stress. During nitrogen starvation, the induction was controlled at the posttranscriptional level. The pexB upstream region contained 245 nucleotides within which sequences approximating the consensus for cyclic AMP receptor protein and integration host factor binding sites were discernible. Deletion of 164 bp of the upstream region, which included these consensus sequences, did not affect starvation-or osmotic stress-mediated induction of pexB but abolished its induction by oxidative stress. The same start site was used in transcription during carbon starvation, osmotic stress, or oxidative stress, suggesting that the pexB promoter can be recognized in vivo by both sigma 38 and sigma 70, depending, presumably, on the presence of appropriate transcriptional factors. The -10 and -35 regions of pexB resembled those of some but not all genes known to be controlled by sigma 38.

    View details for Web of Science ID A1994NU75800013

    View details for PubMedID 8021175



    Transcriptional and translational 'lacZ reporter fusions were constructed to the katF gene, which encodes a putative sigma factor centrally involved in starvation-mediated general resistance in Escherichia coli. Transcription of katF was found to increase ca. twofold after carbon starvation in minimal medium. The protein fusion containing the longest fragment of katF induced ca. eightfold under the same conditions, whereas fusions to shorter segments showed only a twofold increase in expression. The protein fusion was expressed at higher levels in a strain containing a katF::Tn10 mutation, indicating katF autoregulation. The posttranscriptional regulation of katF by starvation did not require a component of the spent minimal medium. katF was also posttranscriptionally regulated during entry into late log phase in complex medium. This induction was coincident with an increase in katE transcription, suggesting that the cellular concentration of KatF directly followed the induction of the katF protein fusion.

    View details for Web of Science ID A1993KU49000036

    View details for PubMedID 8458856



    New computer and statistical methods were used to determine significant direct and inverted repeats in the Escherichia coli contig sequence collection of aggregate 1.6 x 10(6) base-pairs. Eight groups of mostly new structural repeat identities were uncovered. Apart from the high statistical significance of these repeat sequences, there are suggestive relationships of the group matches in terms of neighboring genes, of genomic distributions, of their texts, and of their potentials for secondary structure. Four of these groups are relatively numerous, 11 to 26 members, one is in coding sequences and three are in non-coding. The coding group consists of the ATP-activated transmembrane component of a typical high-affinity protein-binding transport system. One of the non-coding groups consists of a special rho-independent transcription termination signal closely following an operon. The gene neighbors of this group often appear to be involved in some way in processing RNA or DNA. A second non-coding group has, for one or both neighboring genes, a component of a system responding to stress or starvation for some nutrient.

    View details for Web of Science ID A1993KP93900005

    View details for PubMedID 8445651

  • Physiological responses of Lactococcus lactis ML3 to alternating conditions of growth and starvation. Archives of Microbiology Kunji ERS, Ubbink T, Matin A, Poolman B, Konings WN 1993; 159: 372


    The use of glucose starvation to uncouple the production of recombinant beta-galactosidase from cell growth in Escherichia coli was investigated. A lacZ operon fusion to the carbon starvation-inducible cst-1 locus was used to control beta-galactosidase synthesis. beta-Galactosidase induction was observed only under aerobic starvation conditions, and its expression continued for 6 h following the onset of glucose starvation. The cessation of beta-galactosidase expression closely correlated with the exhaustion of acetate, an overflow metabolite of glucose, from the culture medium. Our results suggest the primary role of acetate in cst-1-controlled protein expression is that of an energy source. Using this information, we metered acetate to a glucose-starved culture and produced a metabolically sluggish state, where growth was limited to a low linear rate and production of recombinant beta-galactosidase occurred continuously throughout the experiment. The cst-1 controlled beta-galactosidase synthesis was also induced at low dilution rates in a glucose-limited chemostat, suggesting possible applications to high-density cell systems such as glucose-limited recycle reactors. This work demonstrates that by using an appropriate promoter system and nutrient limitation, growth can be restrained while recombinant protein production is induced and maintained.

    View details for Web of Science ID A1992HX27500010

    View details for PubMedID 18601113