Professional Education


  • Doctor of Philosophy, Institute of Bioorganic Chemistry PAS, Chemistry/Biochemistry (2013)
  • Master of Science/Engineer, Poznań University of Life Sciences, Biotechnology (2008)
  • Master of Science/Engineer, Poznań University of Economics, Commodity of Science (2006)

Stanford Advisors


Lab Affiliations


All Publications


  • Simple alkanoyl acylating agents for reversible RNA functionalization and control. Chemical communications (Cambridge, England) Park, H. S., Kietrys, A. M., Kool, E. T. 2019

    Abstract

    We describe the synthesis and RNA acylation activity of a series of minimalist azidoalkanoyl imidazole reagents, with the aim of functionalizing RNA at 2'-hydroxyl groups at stoichiometric to superstoichiometric levels. We find marked effects of small structural changes on their ability to acylate and be reductively removed, and identify reagents and methods that enable efficient RNA functionalization and control.

    View details for PubMedID 30977472

  • Exceptionally rapid oxime and hydrazone formation promoted by catalytic amine buffers with low toxicity CHEMICAL SCIENCE Larsen, D., Kietrys, A. M., Clark, S. A., Park, H., Ekebergh, A., Kool, E. T. 2018; 9 (23): 5252–59

    View details for DOI 10.1039/c8sc01082j

    View details for Web of Science ID 000435350200016

  • ATP-Linked Chimeric Nucleotide as a Specific Luminescence Reporter of Deoxyuridine Triphosphatase BIOCONJUGATE CHEMISTRY Ji, D., Kietrys, A. M., Lee, Y., Kool, E. T. 2018; 29 (5): 1614–21

    Abstract

    Nucleotide surveillance enzymes play important roles in human health, by monitoring damaged monomers in the nucleotide pool and deactivating them before they are incorporated into chromosomal DNA or disrupt nucleotide metabolism. In particular, deamination of cytosine, leading to uracil in DNA and in the nucleotide pool, can be deleterious, causing DNA damage. The enzyme deoxyuridine triphosphatase (dUTPase) is currently under study as a therapeutic and prognostic target for cancer. Measuring the activity of this enzyme is important both in basic research and in clinical applications involving this pathway, but current methods are nonselective, detecting pyrophosphate, which is produced by many enzymes. Here we describe the design and synthesis of a dUTPase enzyme-specific chimeric dinucleotide (DUAL) that replaces the pyrophosphate leaving group of the native substrate with ATP, enabling sensitive detection via luciferase luminescence signaling. The DUAL probe functions sensitively and selectively to quantify enzyme activities in vitro and in cell lysates. We further report the first measurements of dUTPase activities in eight different cell lines, which are found to vary by a factor of 7-fold. We expect that the new probe can be of considerable utility in research involving this clinically significant enzyme.

    View details for PubMedID 29578692

  • RNA Control by Photoreversible Acylation JOURNAL OF THE AMERICAN CHEMICAL SOCIETY Velema, W. A., Kietrys, A. M., Kool, E. T. 2018; 140 (10): 3491–95

    Abstract

    External photocontrol over RNA function has emerged as a useful tool for studying nucleic acid biology. Most current methods rely on fully synthetic nucleic acids with photocaged nucleobases, limiting application to relatively short synthetic RNAs. Here we report a method to gain photocontrol over RNA by postsynthetic acylation of 2'-hydroxyls with photoprotecting groups. One-step introduction of these groups efficiently blocks hybridization, which is restored after light exposure. Polyacylation (termed cloaking) enables control over a hammerhead ribozyme, illustrating optical control of RNA catalytic function. Use of the new approach on a transcribed 237 nt RNA aptamer demonstrates the utility of this method to switch on RNA folding in a cellular context, and underlines the potential for application in biological studies.

    View details for PubMedID 29474085

  • RNA Cloaking by Reversible Acylation ANGEWANDTE CHEMIE-INTERNATIONAL EDITION Kadina, A., Kietrys, A. M., Kool, E. T. 2018; 57 (12): 3059–63

    Abstract

    We describe a selective and mild chemical approach for controlling RNA hybridization, folding, and enzyme interactions. Reaction of RNAs in aqueous buffer with an azide-substituted acylating agent (100-200 mm) yields several 2'-OH acylations per RNA strand in as little as 10 min. This poly-acylated ("cloaked") RNA is strongly blocked from hybridization with complementary nucleic acids, from cleavage by RNA-processing enzymes, and from folding into active aptamer structures. Importantly, treatment with a water-soluble phosphine triggers a Staudinger reduction of the azide groups, resulting in spontaneous loss of acyl groups ("uncloaking"). This fully restores RNA folding and biochemical activity.

    View details for PubMedID 29370460

    View details for PubMedCentralID PMC5842138

  • Potent and Selective Inhibitors of 8-Oxoguanine DNA Glycosylase JOURNAL OF THE AMERICAN CHEMICAL SOCIETY Tahara, Y., Auld, D., Ji, D., Beharry, A. A., Kietrys, A. M., Wilson, D. L., Jimenez, M., King, D., Nguyen, Z., Kool, E. T. 2018; 140 (6): 2105–14

    Abstract

    The activity of DNA repair enzyme 8-oxoguanine DNA glycosylase (OGG1), which excises oxidized base 8-oxoguanine (8-OG) from DNA, is closely linked to mutagenesis, genotoxicity, cancer, and inflammation. To test the roles of OGG1-mediated repair in these pathways, we have undertaken the development of noncovalent small-molecule inhibitors of the enzyme. Screening of a PubChem-annotated library using a recently developed fluorogenic 8-OG excision assay resulted in multiple validated hit structures, including selected lead hit tetrahydroquinoline 1 (IC50 = 1.7 μM). Optimization of the tetrahydroquinoline scaffold over five regions of the structure ultimately yielded amidobiphenyl compound 41 (SU0268; IC50 = 0.059 μM). SU0268 was confirmed by surface plasmon resonance studies to bind the enzyme both in the absence and in the presence of DNA. The compound SU0268 was shown to be selective for inhibiting OGG1 over multiple repair enzymes, including other base excision repair enzymes, and displayed no toxicity in two human cell lines at 10 μM. Finally, experiments confirm the ability of SU0268 to inhibit OGG1 in HeLa cells, resulting in an increase in accumulation of 8-OG in DNA. The results suggest the compound SU0268 as a potentially useful tool in studies of the role of OGG1 in multiple disease-related pathways.

    View details for PubMedID 29376367

  • Exceptionally rapid oxime and hydrazone formation promoted by catalytic amine buffers with low toxicity. Chemical science Larsen, D., Kietrys, A. M., Clark, S. A., Park, H. S., Ekebergh, A., Kool, E. T. 2018; 9 (23): 5252–59

    Abstract

    Hydrazone and oxime bond formation between α-nucleophiles (e.g. hydrazines, alkoxy-amines) and carbonyl compounds (aldehydes and ketones) is convenient and is widely applied in multiple fields of research. While the reactants are simple, a substantial drawback is the relatively slow reaction at neutral pH. Here we describe a novel molecular strategy for accelerating these reactions, using bifunctional buffer compounds that not only control pH but also catalyze the reaction. The buffers can be employed at pH 5-9 (5-50 mM) and accelerate reactions by several orders of magnitude, yielding second-order rate constants of >10 M-1 s-1. Effective bifunctional amines include 2-(aminomethyl)imidazoles and N,N-dimethylethylenediamine. Unlike previous diaminobenzene catalysts, the new buffer amines are found to have low toxicity to human cells, and can be used to promote reactions in cellular applications.

    View details for PubMedID 29997880

  • Fingerprints of Modified RNA Bases from Deep Sequencing Profiles JOURNAL OF THE AMERICAN CHEMICAL SOCIETY Kietrys, A. M., Velema, W. A., Kool, E. T. 2017; 139 (47): 17074–81

    Abstract

    Posttranscriptional modifications of RNA bases are not only found in many noncoding RNAs but have also recently been identified in coding (messenger) RNAs as well. They require complex and laborious methods to locate, and many still lack methods for localized detection. Here we test the ability of next-generation sequencing (NGS) to detect and distinguish between ten modified bases in synthetic RNAs. We compare ultradeep sequencing patterns of modified bases, including miscoding, insertions and deletions (indels), and truncations, to unmodified bases in the same contexts. The data show widely varied responses to modification, ranging from no response, to high levels of mutations, insertions, deletions, and truncations. The patterns are distinct for several of the modifications, and suggest the future use of ultradeep sequencing as a fingerprinting strategy for locating and identifying modifications in cellular RNAs.

    View details for PubMedID 29111692

  • Chemical and structural effects of base modifications in messenger RNA. Nature Harcourt, E. M., Kietrys, A. M., Kool, E. T. 2017; 541 (7637): 339-346

    Abstract

    A growing number of nucleobase modifications in messenger RNA have been revealed through advances in detection and RNA sequencing. Although some of the biochemical pathways that involve modified bases have been identified, research into the world of RNA modification - the epitranscriptome - is still in an early phase. A variety of chemical tools are being used to characterize base modifications, and the structural effects of known base modifications on RNA pairing, thermodynamics and folding are being determined in relation to their putative biological roles.

    View details for DOI 10.1038/nature21351

    View details for PubMedID 28102265

  • Luminescent Carbon Dot Mimics Assembled on DNA Journal of the American Chemical Society Chan, K., Xu, W., Kwon, H., Kietrys, A. M., Kool, E. T. 2017; 139 (37): 13147–13155

    Abstract

    Nanometer-sized fragments of carbon in the form of multilayer graphene ("carbon dots") have been under highly active study for applications in imaging. While offering advantages of low toxicity and photostability, such nanomaterials are inhomogeneous and have limited wavelengths of emission. Here we address these issues by assembling luminescent aromatic C16-C38 hydrocarbons together on a DNA scaffold in homogeneous, soluble molecular compounds. Monomer deoxyribosides of five different aromatic hydrocarbons were synthesized and assembled into a library of 1296 different tetramer compounds on PEG-polystyrene beads. These were screened for photostability and a range of emission colors using 365 nm excitation, observing visible light (>400 nm) emission. We identified a set of six oligomers (DNA-carbon assemblies, DNA-CAs) with exceptional photostability that emit from 400 to 680 nm in water, with Stokes shifts of up to 110 nm, quantum yields ranging from 0.01 to 0.29, and fluorescence lifetimes from 3 to 42 ns. In addition, several of these DNA-CAs exhibited white emission in aqueous solution. The molecules were used in multispectral cell imaging experiments and were taken up into cells passively. The results expand the range of emission properties that can be achieved in water with all-hydrocarbon chromophores and establish the use of the DNA scaffold to arrange carbon layers in homogeneous, rapidly synthesized assemblies.

    View details for DOI 10.1021/jacs.7b07420

    View details for PubMedCentralID PMC5817913

  • Luminescent Carbon Dot Mimics Assembled on DNA. Journal of the American Chemical Society Chan, K. M., Xu, W., Kwon, H., Kietrys, A. M., Kool, E. T. 2017; 139 (37): 13147–55

    Abstract

    Nanometer-sized fragments of carbon in the form of multilayer graphene ("carbon dots") have been under highly active study for applications in imaging. While offering advantages of low toxicity and photostability, such nanomaterials are inhomogeneous and have limited wavelengths of emission. Here we address these issues by assembling luminescent aromatic C16-C38 hydrocarbons together on a DNA scaffold in homogeneous, soluble molecular compounds. Monomer deoxyribosides of five different aromatic hydrocarbons were synthesized and assembled into a library of 1296 different tetramer compounds on PEG-polystyrene beads. These were screened for photostability and a range of emission colors using 365 nm excitation, observing visible light (>400 nm) emission. We identified a set of six oligomers (DNA-carbon assemblies, DNA-CAs) with exceptional photostability that emit from 400 to 680 nm in water, with Stokes shifts of up to 110 nm, quantum yields ranging from 0.01 to 0.29, and fluorescence lifetimes from 3 to 42 ns. In addition, several of these DNA-CAs exhibited white emission in aqueous solution. The molecules were used in multispectral cell imaging experiments and were taken up into cells passively. The results expand the range of emission properties that can be achieved in water with all-hydrocarbon chromophores and establish the use of the DNA scaffold to arrange carbon layers in homogeneous, rapidly synthesized assemblies.

    View details for PubMedID 28841010

  • Epigenetics: A new methyl mark on messengers. Nature Kietrys, A. M., Kool, E. T. 2016; 530 (7591): 423–24

    View details for PubMedID 26911777

    View details for PubMedCentralID PMC5498786

  • Selection of RNA oligonucleotides that can modulate human dicer activity in vitro. Nucleic acid therapeutics Tyczewska, A., Kurzyńska-Kokorniak, A., Koralewska, N., Szopa, A., Kietrys, A. M., Wrzesiński, J., Twardowski, T., Figlerowicz, M. 2011; 21 (5): 333–46

    Abstract

    Human ribonuclease Dicer is an enzyme that excises small regulatory RNAs from perfectly or partially double-stranded RNA precursors. Although Dicer substrates and products have already been quite well characterized, our knowledge about cellular factors regulating the activity of this enzyme is still limited. To learn more about this problem, we attempted to determine whether RNA could function not only as a Dicer substrate but also as its regulator. To this end, we applied an in vitro selection method. We identified 120 RNA oligomers binding human Dicer. Sixteen of them were subjected to more detailed in vitro studies. We found that 6 out of 16 oligomers affected Dicer ability to digest pre-microRNAs (miRNAs), although most of them were cleaved by this enzyme. For the 6 most active oligomers the putative mechanism of Dicer inhibition was determined. Three oligomers were classified as typical competitive inhibitors and one as an allosteric inhibitor. The remaining 2 oligomers acted as selective inhibitors. They affected the production of 1 miRNA, whereas the formation of other miRNAs was hardly influenced. In general, the data obtained suggest that one can modulate the generation of specific miRNAs by using RNA oligomers. Moreover, we found that sequences similar to those of the selected oligomers can be found within the molecules composing human transcriptome.

    View details for DOI 10.1089/nat.2011.0304

    View details for PubMedID 22004415

  • Antisense oligonucleotides targeting universally conserved 26S rRNA domains of plant ribosomes at different steps of polypeptide elongation. Oligonucleotides Bakowska-Zywicka, K., Kietrys, A. M., Twardowski, T. 2008; 18 (2): 175–86

    Abstract

    A ribosome undergoes significant conformational changes during elongation of polypeptide chain that are correlated with structural changes of rRNAs. We tested nine different antisense oligodeoxynucleotides complementary to the selected, highly conserved sequences of Lupinus luteus 26S rRNA that are engaged in the interactions with tRNA molecules. The ribosomes were converted either to pre- or to posttranslocational states, with or without prehybridized oligonucleotides, using tRNA or mini-tRNA molecules. The activity of those ribosomes was tested via the so-called binding assay. We observed well-defined structural changes of ribosome's conformation during different steps of the elongation cycle of protein biosynthesis. In this article, we present that (i) before and after translocation, fragments of domain V between helices H70/H71 and H74/H89 do not have to interact with nucleotides 72-76 of the acceptor arm of A-site tRNA; (ii) helix H69 does not have to interact with DHU arm of tRNA in positions 25 and 26 after forming the peptide bond, but before translocation; (iii) helices H69 and H70 interact weakly with nucleotides 11, 12, 25, and 26 of A-site tRNA before forming a peptide bond in the ribosome; (iv) interactions between helices H80, H93 and single-stranded region between helices H92 and H93 and CCAend of P-site tRNA are necessary at all steps of elongation cycle; and (v) before and after translocation, helix H89 does not have to interact with nucleotides in positions 64-65 and 50-53 of A-site tRNA TPsiC arm.

    View details for DOI 10.1089/oli.2008.0127

    View details for PubMedID 18637734