Proceedings of the National Academy of Sciences of the United States of America, 13 February 2001, Vol.98(4), pp.1571-1576
Unidirectional proton transport in bacteriorhodopsin is enforced by the switching machinery of the active site. Threonine 89 is located in this region, with its O-H group forming a hydrogen bond with Asp-85, the acceptor for proton transfer from the Schiff base of the retinal chromophore. Previous IR spectroscopy of [3- 18 O]threonine-labeled bacteriorhodopsin showed that the hydrogen bond of the O-D group of Thr-89 in D 2O is strengthened in the K photocycle intermediate. Here, we show that the strength and orientation of this hydrogen bond remains unchanged in the L intermediate and through the M intermediate. Furthermore, a strong interaction between Asp-85 and the O-H (O-D) group of Thr-89 in M is indicated by a shift in the C=O stretching vibration of the former because of 18 O substitution in the latter. Thus, the strong hydrogen bond between Asp-85 and Thr-89 in K persists through M, contrary to structural models based on x-ray crystallography of the photocycle intermediates. We propose that, upon photoisomerization of the chromophore, Thr-89 forms a tight, persistent complex with one of the side-chain oxygens of Asp-85 and is thereby precluded from participating in the switching process. On the other hand, the loss of hydrogen bonding at the other oxygen of Asp-85 in M may be related to the switching event.
Physical sciences -- Chemistry -- Chemical bonding ; Physical sciences -- Physics -- Matter ; Physical sciences -- Chemistry -- Chemical elements ; Physical sciences -- Chemistry -- Chemical compounds ; Physical sciences -- Physics -- Microphysics ; Applied sciences -- Laboratory techniques -- Spectroscopy ; Physical sciences -- Physics -- Microphysics ; Physical sciences -- Physics -- Microphysics ; Physical sciences -- Physics -- Mechanics ; Physical sciences -- Chemistry -- Chemical reactions
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