Professional Education


  • Vordiplom, Technische Universitat Berlin (2007)
  • Diplom, Technische Universitat Berlin (2009)
  • Doctor of Philosophy, Technische Universitat Berlin (2013)

Stanford Advisors


All Publications


  • Monitoring the Transmembrane Proton Gradient Generated by Cytochrome bo(3) in Tethered Bilayer Lipid Membranes Using SEIRA Spectroscopy JOURNAL OF PHYSICAL CHEMISTRY B Wiebalck, S., Kozuch, J., Forbrig, E., Tzschucke, C. C., Jeuken, L. J., Hildebrandt, P. 2016; 120 (9): 2249-2256

    Abstract

    Membrane proteins act as biocatalysts or ion/proton pumps to convert and store energy from ubiquitous environmental sources. Interfacing these proteins to electrodes allows utilizing the energy for enzymatic biofuel cells or other auspicious biotechnological applications. To optimize the efficiency of these devices, appropriate membrane models are required that ensure structural and functional integrity of the embedded enzymes and provide structural insight. We present a spectroelectrochemical surface-enhanced infrared absorption (SEIRA) and electrical impedance spectroscopy (EIS) study of the bacterial respiratory ubiquinol/cytochrome bo3 (cyt bo3) couple incorporated into a tethered bilayer lipid membrane (tBLM). Here, we employed a new lipid tether (WK3SH, dihydrocholesteryl (2-(2-(2-ethoxy)ethoxy)ethanethiol), which was synthesized using a three-step procedure with very good yield and allowed measuring IR spectra without significant spectral interference of the tBLM. The functional integrity of the incorporated cyt bo3 was demonstrated by monitoring the enzymatic O2 reduction current and the formation of the transmembrane proton gradient. Based on a SEIRA-spectroscopic redox titration, a shift of the pH-dependent redox potential of the ubiquinones under turnover conditions was correlated with an alkalinization of the submembrane reservoir by +0.8 pH units. This study demonstrates the high potential of tBLMs and the SEIRA spectroscopic approach to study bioenergetic processes.

    View details for DOI 10.1021/acs.jpcb.6b01435

    View details for Web of Science ID 000372042000020

    View details for PubMedID 26898921

  • Voltage-dependent structural changes of the membrane-bound anion channel hVDAC1 probed by SEIRA and electrochemical impedance spectroscopy PHYSICAL CHEMISTRY CHEMICAL PHYSICS Kozuch, J., Weichbrodt, C., Millo, D., Giller, K., Becker, S., Hildebrandt, P., Steinem, C. 2014; 16 (20): 9546-9555

    Abstract

    The voltage-dependent anion channel (VDAC) is a transmembrane protein that regulates the transfer of metabolites between the cytosol and the mitochondrium. Opening and partial closing of the channel is known to be driven by the transmembrane potential via a mechanism that is not fully understood. In this work, we employed a spectroelectrochemical approach to probe the voltage-induced molecular structure changes of human VDAC1 (hVDAC1) embedded in a tethered bilayer lipid membrane on a nanostructured Au electrode. The model membrane consisted of a mixed self-assembled monolayer of 6-mercaptohexanol and (cholesterylpolyethylenoxy)thiol, followed by the deposition of 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine vesicles including hVDAC1. The stepwise assembly of the model membrane and the incorporation of hVDAC1 were monitored by surface enhanced infrared absorption and electrochemical impedance spectroscopy. Difference spectra allowed for identifying the spectral changes which may be associated with the transition from the open to the "closed" states by shifting the potential above or below the transmembrane potential determined to be ca. 0.0 V vs. the open circuit potential. These spectral changes were interpreted on the basis of the orientation- and distance-dependent IR enhancement and indicate alterations of the inclination angle of the β-strands as crucial molecular events, reflecting an expansion or contraction of the β-barrel pore. These protein structural changes that do not confirm nor exclude the reorientation of the α-helix are either directly induced by the electric field or a consequence of a potential-dependent repulsion or attraction of the bilayer.

    View details for DOI 10.1039/c4cp00167b

    View details for Web of Science ID 000335818600039

    View details for PubMedID 24728177

  • Combined Electrochemistry and Surface-Enhanced Infrared Absorption Spectroscopy of Gramicidin A Incorporated into Tethered Bilayer Lipid Membranes ANGEWANDTE CHEMIE-INTERNATIONAL EDITION Kozuch, J., Steinem, C., Hildebrandt, P., Millo, D. 2012; 51 (32): 8114-8117

    View details for DOI 10.1002/anie.201203214

    View details for Web of Science ID 000307215600054

    View details for PubMedID 22865570

  • Switchable Redox Chemistry of the Hexameric Tyrosine-Coordinated Heme Protein JOURNAL OF PHYSICAL CHEMISTRY B Kielb, P., Utesch, T., Kozuch, J., Jeoung, J., Dobbek, H., Mroginski, M. A., Hildebrandt, P., Weidinger, I. 2017; 121 (16): 3955-3964

    Abstract

    Hexameric tyrosine-coordinated heme protein HTHP from Silicibacter pomeroyi has been shown to exhibit peroxidase- and catalase-like activity. In the present study, detailed spectroscopic and electrochemical investigations were performed to analyze the redox properties and active site structure of HTHP. Potentiometric titration of HTHP in solution revealed a single redox transition at -0.54 V (vs Ag/AgCl), indicating six structurally identical tyrosine coordinates hemes. Cyclic voltammetry (CV) of immobilized HTHP afforded a distinctly more positive redox potential (-0.17 V) but failed to detect a transition at -0.54 V. Conversely, surface enhanced RR (SERR) spectroscopy provided evidence for both high- and low-potential transitions and for a partial loss of heme in the reduced state. The high-potential CV-active redox transition is attributed to the hemes of the barrel-shaped HTHP in a wheel-like orientation on the surface. Supported by coarse-grained simulations and SERR spectroscopy, the majority of HTHP is concluded to adopt a reverse-disc orientation, accounting for the low-potential transition. In view of the striking similarity of HTHP to the heme carriers HasA or HmbR regarding redox potential, Fe-Tyr ligation, and heme release, we propose heme transport as an alternative or additional function.

    View details for DOI 10.1021/acs.jpcb.7b01286

    View details for Web of Science ID 000400534200006

    View details for PubMedID 28383909

  • Calculating average surface enhancement factors of randomly nanostructured electrodes by a combination of SERS and impedance spectroscopy PHYSICAL CHEMISTRY CHEMICAL PHYSICS Kozuch, J., Petrusch, N., Gkogkou, D., Gernert, U., Weidinger, I. M. 2015; 17 (33): 21220-21225

    Abstract

    Polyhedron Ag nanostructures were created on top of a polished Au electrode via step-wise electrodeposition and tested as substrates for SERS spectroscopy. Average Raman enhancement factors were derived by combining SERS measurements with electrochemical impedance spectroscopy (EIS), which is able to determine the electroactive surface area of a randomly nanostructured surface. Depending on the deposition step an alternating increase and decrease of surface area was observed while the SERS intensity showed a clear maximum for the first deposition cycle. SEM pictures reveal the formation of Ag polyhedrons that are randomly dispersed on the Au surface. Furthermore the presence of a sub nanostructure on top of the polyhedron after the first deposition cycle is observed which becomes smoother after subsequent deposition cycles. Correlating the SEM pictures with SERS and EIS measurements it is concluded that the coral-like sub nanostructure is dominating the enhancement factor while the polyhedron structure itself only plays a minor role for electromagnetic field enhancement.

    View details for DOI 10.1039/c4cp05015k

    View details for Web of Science ID 000359596600021

    View details for PubMedID 25599525

  • 2nd coordination sphere controlled electron transfer of iron hangman complexes on electrodes probed by surface enhanced vibrational spectroscopy CHEMICAL SCIENCE Ly, H. K., Wrzolek, P., Heidary, N., Goetz, R., Horch, M., Kozuch, J., Schwalbe, M., Weidinger, I. M. 2015; 6 (12): 6999-7007

    View details for DOI 10.1039/c5sc02560e

    View details for Web of Science ID 000365225300037

  • Potential-Dependent Surface-Enhanced Resonance Raman Spectroscopy at Nanostructured TiO2 : A Case Study on Cytochrome b(5) SMALL Han, X. X., Koehler, C., Kozuch, J., Kuhlmann, U., Paasche, L., Sivanesan, A., Weidinger, I. M., Hildebrandt, P. 2013; 9 (24): 4175-4181

    Abstract

    Nanostructured titanium dioxide (TiO2 ) electrodes, prepared by anodization of titanium, are employed to probe the electron-transfer process of cytochrome b5 (cyt b5 ) by surface-enhanced resonance Raman (SERR) spectroscopy. Concomitant with the increased nanoscopic surface roughness of TiO2 , achieved by raising the anodization voltage from 10 to 20 V, the enhancement factor increases from 2.4 to 8.6, which is rationalized by calculations of the electric field enhancement. Cyt b5 is immobilized on TiO2 under preservation of its native structure but it displays a non-ideal redox behavior due to the limited conductivity of the electrode material. The electron-transfer efficiency which depends on the crystalline phase of TiO2 has to be improved by appropriate doping for applications in bioelectrochemistry.

    View details for DOI 10.1002/smll.201301070

    View details for Web of Science ID 000328453300012

    View details for PubMedID 23861351

  • Resonance Raman Characterization of the Ammonia-Generated Oxo Intermediate of Cytochrome c Oxidase from Paracoccus denitrificans BIOCHEMISTRY Kozuch, J., von der Hocht, I., Hilbers, F., Michel, H., Weidinger, I. M. 2013; 52 (36): 6197-6202

    Abstract

    A novel oxo state of cytochrome c oxidase from Paracoccus denitrificans generated by successive addition of excess H2O2 and ammonia was investigated using resonance Raman (RR) spectroscopy. Addition of ammonia to the H2O2-generated artificial F state resulted in an upshift of the oxoferryl stretching vibration from 790 to 796 cm(-1), indicating that ammonia influences ligation of the heme-bound oxygen in the binuclear center. Concomitantly performed RR measurements in the high-frequency region between 1300 and 1700 cm(-1) showed a high-spin to low-spin transition of heme a3 upon generation of the F state that was not altered by addition of ammonia. Removal of H2O2 by addition of catalase resulted in the disappearance of the oxoferryl stretching vibration and major back transformation of heme a3 into the high-spin state. The ratio of high-spin to low-spin states was identical for intermediates created with and without ammonia, leading to the conclusion that ammonia does not interact directly with heme a3. Only for the ammonia-created state was a band at 612 nm observed in the UV-visible difference spectrum that was shifted to 608 nm after addition of catalase. Our results support the hypothesis by von der Hocht et al. [von der Hocht, I., et al. (2011) Proc. Natl. Acad. Sci. U.S.A. 108, 3964-3969] that addition of ammonia creates a novel oxo intermediate state called PN where ammonia binds to CuB once the oxo intermediate F state has been formed.

    View details for DOI 10.1021/bi400535m

    View details for Web of Science ID 000330099900007

    View details for PubMedID 23914722

  • Tailored silica coated Ag nanoparticles for non-invasive surface enhanced Raman spectroscopy of biomolecular targets RSC ADVANCES Sivanesan, A., Kozuch, J., Ly, H. K., Kalaivani, G., Fischer, A., Weidinger, I. M. 2012; 2 (3): 805-808

    View details for DOI 10.1039/c1ra00781e

    View details for Web of Science ID 000299177000009

  • Functionalized Ag nanoparticles with tunable optical properties for selective protein analysis CHEMICAL COMMUNICATIONS Sivanesan, A., Ly, H. K., Kozuch, J., Sezer, M., Kuhlmann, U., Fischer, A., Weidinger, I. M. 2011; 47 (12): 3553-3555

    Abstract

    We present a preparation procedure for small sized biocompatibly coated Ag nanoparticles with tunable surface plasmon resonances. The conditions were optimised with respect to the resonance Raman signal enhancement of heme proteins and to the preservation of the native protein structure.

    View details for DOI 10.1039/c0cc05058j

    View details for Web of Science ID 000288085600065

    View details for PubMedID 21321696