Complete Proton Transfer Cycle in GFP and Its T203V and S205V Mutants |
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| Authors: | Dr Sergey P Laptenok Prof Andras Lukacs Agnieszka Gil Dr Richard Brust Dr Igor V Sazanovich Dr Gregory M Greetham Prof Peter J Tonge Prof Stephen R Meech |
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| Institution: | 1. School of Chemistry, University of East Anglia, Norwich, NR4 7TJ (UK);2. Department of Biophysics, Medical School, Szigeti str. 12, 7624 Pécs (Hungary);3. Department of Chemistry, Stony Brook University, Stony Brook, NY 11794‐3400 (USA);4. The Scripps Research Institute, 130 Scripps Way, Jupiter, FL 33458 (USA);5. Central Laser Facility, Research Complex at Harwell, Harwell Science and Innovation Campus, Didcot, Oxon OX11 0QX (UK) |
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| Abstract: | Proton transfer is critical in many important biochemical reactions. The unique three‐step excited‐state proton transfer in avGFP allows observations of protein proton transport in real‐time. In this work we exploit femtosecond to microsecond transient IR spectroscopy to record, in D2O, the complete proton transfer photocycle of avGFP, and two mutants (T203V and S205V) which modify the structure of the proton wire. Striking differences and similarities are observed among the three mutants yielding novel information on proton transfer mechanism, rates, isotope effects, H‐bond strength and proton wire stability. These data provide a detailed picture of the dynamics of long‐range proton transfer in a protein against which calculations may be compared. |
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| Keywords: | green fluorescent protein (GFP) IR spectroscopy kinetic isotope effect proton transfer ultrafast spectroscopy |
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