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Project B4

Hydrogen bonding network in channelrhodopsins analyzed by site-directed labeling and EPR spectroscopy

Principal Investigators: Prof. Dr. Robert Bittl (FU), Dr. Ramona Schlesinger (FU)

Conformational changes and alterations in the hydrogen bonding network of channelrhodopsin will be investigated by site-directed labeling and electron paramagnetic resonance (EPR) spectroscopy. Constructs will be generated where the native cysteine residues are reduced to a minimum and new cysteines for spin labeling are introduced to detect changes in polarity and hydrogen bonding by virtue of the sensitivity of the spin-label EPR parameters. Moreover the project aims at identifying and analyzing protein conformational changes essential for understanding protonation dynamics using pulsed electron-electron double resonance on doubly spin-labeled constructs.

Publications

2017 - 2020

Ehrenberg, D., Krause, N., Saita, M., Bamann, C., Kar, R.K., Hoffmann, K., Heinrich, D., Schapiro, I.Heberle, J., and Schlesinger, R. (2019) Atomistic insight into the role of threonine 127 in the functional mechanism of channelrhodopsin-2. Appl. Sci., 9 (22), 4905; https://doi.org/10.3390/app9224905.

Harris N.J., Reading E., Ataka K., Grzegorzewski L., Charalambous K., Liu X., Schlesinger R., Heberle J., and Booth P.J. (2017). Structure formation during translocon-unassisted co-translational membrane protein folding. Scientific Reports 7: 8021; doi: 10.1038/s41598-017-08522-9

Kovácsová, G., Grunbein, M.L., Kloos, M., Barends, T.R.M., Schlesinger, R., Heberle, J., Kabsch, W., Shoeman, R.L., Doak, R.B., and Schlichting, I. (2017). Viscous hydrophilic injection matrices for serial crystallography. IUCrJ 4, 400-410; doi: 10.1107/S2052252517005140

Kruse, F., Nguyen, A.D., Dragelj, J., Schlesinger, R., Heberle, J., Mroginski, M.A., and Weidinger, I.M. (2020). Characterisation of the Cyanate Inhibited State of Cytochrome c Oxidase. Sci. Rep. 10, 3863; doi: 10.1038/s41598-020-60801-0

Lorenz-Fonfria V.A., Saita M., Lazarova T., Schlesinger R., and Heberle J. (2017). pH-sensitive vibrational probe reveals a cytoplasmic protonated cluster in bacteriorhodopsin. Proceedings of the National Academy of Sciences of the United States of America 114, E10909-E10918; doi: 10.1073/pnas.1707993114

Nass Kovacs, G., Colletier, J.-P., Grünbein, M. L., Yang, Y., Stensitzki, T., Batyuk, A., Carbajo, S., Doak, R. B., Ehrenberg, D., Foucar, L., Gasper, R., Gorel, A., Hilpert, M., Kloos, M., Koglin, J. E., Reinstein, J., Roome, C. M., Schlesinger, R., Seaberg, M., Shoeman, R. L., Stricker, M., Boutet, S., Haacke, S., Heberle, J., Heyne, K., Domratcheva, T., Barends, T. R. M., and Schlichting, I.(2019). Three-dimensional view of ultrafast dynamics in photoexcited bacteriorhodopsin. Nature Communications 10, Article Number 3177. doi: 10.1038/s41467-019-10758-0

Pfitzner, E., Seki, H., Schlesinger, R., Ataka, K., and Heberle, J. (2018). Disc Antenna Enhanced Infrared Spectroscopy: From Self-Assembled Monolayers to Membrane Proteins. ACS Sens, 3 (5), pp 984–991, doi:10.1021/acssensors.8b00139.

Saita, M., Pranga-Sellnau, F., Resler, T., Schlesinger, R., Heberle, J., and Lorenz-Fonfria, V.A. (2018). Photoexcitation of the P4(480) State Induces a Secondary Photocycle That Potentially Desensitizes Channelrhodopsin-2. J. Am. Chem. Soc. 140, 9899-9903; doi: 10.1021/jacs.8b03931

Sezer, M., Woelke, A.-L., Knapp, E.W., Schlesinger, R., Mroginski, M.A., Weidinger, I.M. (2017). Redox induced protonation of heme propionates in cytochrome c oxidase: Insights from surface enhanced resonance Raman spectroscopy and QM/MM calculations. Biochimica et Biophysica Acta (BBA) - Bioenergetics 1858, 103-108; doi: 10.1016/j.bbabio.2016.10.009

Stensitzki, T., Adam, S., Schlesinger, R., Schapiro, I., and Heyne, K. (2020). Ultrafast Backbone Protonation in Channelrhodopsin-1 Captured by Polarization Resolved Fs Vis-pump-IR-Probe Spectroscopy and Computational Methods. Molecules 25, 848; doi: 10.3390/molecules25040848

Weber A. M., Kaiser, J., Ziegler, T., Pilsl, S.,, Renzl, C., Sixt, L., Pietruschka, G., Moniot, S., Kakoti, A., Juraschitz, M., Schrottke, S.,Lledo Bryant, L., Steegborn, C., Bittl, R., Mayer, G., and Möglich, A. (2019). A blue light receptor that mediates RNA binding and translational regulation. Nature Chemical Biology, doi: 10.1038/s41589-019-0346-y

2013 - 2016

Krause, N., Engelhard, C., Heberle, J., Schlesinger, R., and Bittl, R. (2013). Structural differences between the closed and open states of channelrhodopsin-2 as observed by EPR spectroscopy. FEBS Letters 587, 3309-3313.

Lórenz-Fonfría, V.A., Bamann, C., Resler, T., Schlesinger, R., Bamberg, E., and Heberle, J. (2015). Temporal evolution of helix hydration in a light-gated ion channel correlates with ion conductance. Proceedings of the National Academy of Sciences of the United States of America 112, E5796-E5804.

Lórenz-Fonfría, V.A., Muders, V., Schlesinger, R., and Heberle, J. (2014). Changes in the hydrogen-bonding strength of internal water molecules and cysteine residues in the conductive state of channelrhodopsin-1. Journal of Chemical Physics 141, 22D507.

Lorenz-Fonfria, V.A., Resler, T., Krause, N., Nack, M., Gossing, M., von Mollard, G.F., Bamann, C., Bamberg, E., Schlesinger, R., and Heberle, J. (2013). Transient protonation changes in channelrhodopsin-2 and their relevance to channel gating. Proceedings of the National Academy of Sciences of the United States of America 110, E1273-E1281.

Lórenz-Fonfría, V.A., Schultz, B.-J., Resler, T., Schlesinger, R., Bamann, C., Bamberg, E., and Heberle, J. (2015). Pre-Gating Conformational Changes in the ChETA Variant of Channelrhodopsin-2 Monitored by Nanosecond IR Spectroscopy. Journal of the American Chemical Society 137, 1850-1861.

Muders, V., Kerruth, S., Lorenz-Fonfria, V.A., Bamann, C., Heberle, J., and Schlesinger, R. (2014). Resonance Raman and FTIR spectroscopic characterization of the closed and open states of channelrhodopsin-1. FEBS Letters 588, 2301-2306.

Resler, T., Schultz, B.-J., Lorenz-Fonfria, V.A., Schlesinger, R., and Heberle, J. (2015). Kinetic and Vibrational Isotope Effects of Proton Transfer Reactions in Channelrhodopsin-2. Biophysical Journal 109, 287-297.

Schnedermann, C., Muders, V., Ehrenberg, D., Schlesinger, R., Kukura, P., and Heberle, J. (2016). Vibronic Dynamics of the Ultrafast all-trans to 13-cis Photoisomerization of Retinal in Channelrhodopsin-1. Journal of the American Chemical Society 138, 4757-4762.

Sezer, M., Kielb, P., Kuhlmann, U., Mohrmann, H., Schulz, C., Heinrich, D., Schlesinger, R., Heberle, J., and Weidinger, I.M. (2015). Surface Enhanced Resonance Raman Spectroscopy Reveals Potential Induced Redox and Conformational Changes of Cytochrome c Oxidase on Electrodes. Journal of Physical Chemistry B 119, 9586-9591.

Stensitzki, T., Muders, V., Schlesinger, R., Heberle, J., and Heyne, K. (2015). The primary photoreaction of channelrhodopsin-1: Wavelength dependent photoreactions induced by ground-state heterogeneity. Frontiers in Molecular Biosciences 2, 41.

Stensitzki, T., Yang, Y., Muders, V., Schlesinger, R., Heberle, J., and Heyne, K. (2016). Femtosecond infrared spectroscopy of channelrhodopsin-1 chromophore isomerization. Structural Dynamics 3, 043208.

Volz, P., Krause, N., Balke, J., Schneider, C., Walter, M., Schneider, F., Schlesinger, R., and Alexiev, U. (2016). Light and pH-induced changes in structure and accessibility of transmembrane helix B and its immediate environment in Channelrhodopsin-2. The Journal of Biological Chemistry 291, 17382-17393.