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Proton Travel in Green Fluorescent Protein
Volkhard Helms and Wei Gu
Abstract Green fluorescence protein (GFP) wild type and some of its mutants
undergo excited state proton transfer between the chromophore and the nearby
Glu222 residue. This process has been covered in detail in the chapter written by
Stephen Meech. Apart from this ultrafast photochemical reaction, multiple other
proton-transfer processes take place in the GFP protein matrix, and these will be
covered in this chapter. For example, proton exchange between the chromophore
and the nearby bulk solvent may occur via His148 that is located in hydrogen-
bonding distance from the chromophore and provides direct access to the bulk
solvent. Moreover, two extended proton-transfer wires including titratable residues
as well as a number of buried water molecules connect the chromophore to the
protein surface. Based on a recent high-resolution X-ray structure of GFP, all
titratable groups of the protein could be placed in one of these two large hydro-
gen-bonding clusters, suggesting that a multitude of proton-transfer processes can
occur in the GFP matrix at any moment in time. While it is quite likely that similar
proton pathways also exist in other soluble and membrane proteins, they are much
harder to study. GFP is an exciting model system for monitoring those processes as
they often directly affect the chromophore photophysics. The dynamics of proton
exchange inside the GFP barrel and with bulk solvent has thus been characterized
by fluorescence correlation spectroscopy (FCS) of the chromophore fluorescence
and by pH-jump experiments. These studies showed that the autocorrelation of the
chromophore fluorescence is affected either by pH-independent processes on
microsecond to millisecond time scales or by pH-dependent processes on similar
time scales. The former ones are likely proton equilibria occurring within the GFP
barrel, and the latter ones are likely exchange processes with the solvent. Biomo-
lecular simulation methods are now being developed, which will soon allow
accessing such time scales by computational means. Then, we will hopefully be
able to connect the spectroscopic findings with dynamic atomistic simulations of
proton-transfer dynamics.
V. Helms ( * ) and W. Gu
Center for Bioinformatics, Saarland University, Campus C7.1, 66123 Saarbruecken, Germany
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