Primary Reaction Dynamics of Proteorhodopsin Mutant D97N Observed by Femtosecond Infrared and Visible Spectroscopy† |
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| Authors: | Mirka‐Kristin Verhoefen Karsten Neumann Ingrid Weber Clemens Glaubitz Josef Wachtveitl |
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| Institution: | 1. Institute of Physical and Theoretical Chemistry/Institute of Biophysics, Johann Wolfgang Goethe‐University Frankfurt, Frankfurt am Main, Germany;2. Institute of Biophysical Chemistry, Johann Wolfgang Goethe‐University Frankfurt, Frankfurt am Main, Germany |
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| Abstract: | Femtosecond time‐resolved spectroscopy in the visible and IR range was utilized to study the primary reaction dynamics of the proteorhodopsin (PR) D97N mutant in comparison with wild type PR at different pH values. The analysis of the data obtained in the mid‐IR closely resembles the results for wild type PR. The observation of the first ground state intermediate K is initially obscured by a complex reaction scheme of vibrational relaxation and heating effects, but its spectral signature clearly emerges at long delay times. In the visible range, a biexponential decay of the excited state within 30 ps and the formation of the K photoproduct is observed. The decay time constants derived for the D97N mutant in D2O are slightly larger than in H2O due to H/D exchange. This kinetic isotope effect is even less pronounced than for wild type PR at pH 6. These results support the current notion of a pH dependent hydrogen bonding network in the retinal binding pocket of PR and a weaker interaction between the retinal Schiff base and the counter ion complex compared to bacteriorhodopsin. |
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