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Principal Investigator

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P5  Werner Mäntele
Professor and Director

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Institute of Biophysics
Goethe University Frankfurt a.M.
Max-von-Laue-Str.1
60438 Frankfurt am Main, Germany


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P5 Structural properties, activation mechanisms and transport processes in betaine transporter BetP and Na+/H+ antiporters NhaA/MjNhaP1 investigated by FT-IR and CD spectroscopy

 

In this project we investigate structural properties, mechanisms of activation and molecular details of transport for secondary transporters. We focus on the Na+/H+ antiporters NhaA from E. coli and MjNhaP1 from Methanococcus jannaschii and on the betaine transporter protein BetP from Corynebacterium glutamicum. These transporters are activated by pH (NhaA and MjNhaP1) and ionic strength (BetP). Although structural information is available, the details of pH or ionic strength sensing, transduction of this information to the transport site, as well as details of the transport mechanism are still unclear.

We use Fourier transform infrared (FT-IR) spectroscopic techniques to obtain structural and functional information on these transporters. Steady-state IR spectroscopy is used to obtain secondary structure information for different functional states of the proteins, and the kinetics and extent of 1H/2H exchange is used to study the respective accessible surface. A perfusion technique based on evanescent wave spectroscopy is used for precise reaction control and functional analysis. For example, inactive-to-active switching can be reversibly induced and monitored spectroscopically in situ. Most of the IR spectroscopic experiments are complemented by CD spectroscopy.

In order to identiy the role of individual amino acids or protein parts as pH sensors or osmosensors, we study site-directed mutants. For an understanding of the structural and functional role of lipid-protein-interactions, we use selected protein and lipid vibrational modes as probes. Together with the available crystal structure of NhaA and the structural data from BetP, these functional investigations will lead to an understanding of pH sensing, osmosensing, signal transducing and transport mechanisms on a molecular level.

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Publications

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Verhoefen, M.K., Schäfer, G., Shastri, S. Weber, I., Glaubitz, C., Mäntele, W. and Wachtveitl, J. (2011) Low temperature FTIR spectroscopy provides new insights in the pH-dependent proton pathway of proteorhodopsin. Biochim Biophys Acta (Bioenergetics) 1807, 1583-1590.

Korkmaz-Özkan, F., Köster, S., Kühlbrandt, W., Mäntele, W. and Yildiz, Ö. (2010) Correlation between the OmpG secondary structure and its pH dependent alterations monitored by FTIR. J Mol Biol 401,56-67.

Stolz, M., Lewitzky, E., Bergbauer, R., Mäntele, W., Grell, E. and Barth, A. (2009) Structural Changes in the Catalytic Cycle of the Na+,K+-ATPase Studied by Infrared Spectroscopy. Biophys J 96, 3433-3442.

Schäfer, G., Shastri, S., Verhoefen, M. K., Glaubitz, C., Wachtveitl, J. and Mäntele, W. (2009) Characterizing the Structure and Photocycle of PR 2D Crystals with CD and FTIR Spectroscopy. Photochem and Photobiol 85, 529-534.

Džafić, E., Klein, O., Screpanti, E., Hunte, C. and Mäntele, W. (2009) Flexibility and dynamics of NhaA Na+/H+-antiporter of Escherichia coli studied by Fourier transform infrared spectroscopy. Spectrochim Acta A 72, 102-109.

Džafić, E., Klein, O., Goswami, P., Kühlbrandt, W. and Mäntele, W. (2009) Infrared spectroscopic study of the structural and functional properties of the Na+/H+ antiporter MjNhaP1 from Methanococcus jannaschii. BBA-Bioenergetics 1787, 730-737.

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Collaborations

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Wachtveitl (P12), Ziegler (P4), Kühlbrandt (P1), Michel (P3), Fendler/Bamberg (P10), Dötsch/Bernhard (P2), Glaubitz (P6), Abele (P9), Prisner (P7)