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Biophysics of Proteins at Surfaces: Assembly, Activation, Signaling
Thursday Speaker Abstracts
Germanium Catches Proteins in Action
Jonas Schartner
, Jörn Güldenhaupt, Konstantin Gavriljuk, Andreas Nabers, Klaus Gerwert,
Carsten Kötting.
Biology & Biotechnology, Bochum, Germany.
The attenuated total reflection fourier transform infrared (ATR-FTIR) spectroscopy allows a
detailed analysis of surface attached molecules, including their secondary structure, reaction
mechanism, orientation, and interaction with small molecules.[1] This technique reveals
vibrational changes in the attached molecules. We recently developed a universal immobilization
technique for the specific immobilization of N-Ras and Photosystem I on a silane modified
germanium surface.[1]
We now present a new approach employing thiol chemistry on germanium.[2] On one hand
germanium crystals provide a great signal-to-noise ratio in ATR-FTIR. On the other hand protein
immobilization via thiol chemistry is well-established because it is standard for modifications of
gold surfaces e.g. in surface plasmon resonance. Here we combine the best of both worlds and
report on germanium surface functionalization with different thiols which allowed for specific
immobilization of histidine-tagged proteins with over 99% specific binding. The great
advantages of using thiols in comparison with silanes are that a huge variety of thiols with
functional groups is commercially available and the monolayer stability is very high. Nativity of
protein folding was confirmed by secondary structure analysis. Stimulus induced difference
spectra were obtained for immobilized Channelrhodopsin 2, the small GTPase N-Ras and the
phosphocholine-transferase AnkX, which demonstrated protein function at atomic level.[3]
To further improve the S/N ratio the establishment of a 3D-surface was achieved by tethering
dextran-polymers to germanium. Proteins were immobilized in multilayers with a distance of
about 9 nm as shown for mCherry and GFP.[4] The difference signal was increased by more than
factor three when tris-ANTA was employed for catching proteins in action.
References
1: Schartner J. et al., JACS, 2013, 135, 4079-4087
2: Han, S. et al. JACS, 2001, 123, 2422–2425
3: Schartner J., et al., ChemBioChem, 2014, 2529-2534
4: Schartner J, et al., under review, 2015