J. Biol. Chem., Vol. 269, Issue 20, 14353-14354, May, 1994
Lowering the intrinsic pKa of the chromophore's Schiff base can restore its light-induced deprotonation in the inactive Tyr-57-->Asn mutant of bacteriorhodopsin
R Govindjee, S Balashov, T Ebrey, D Oesterhelt, G Steinberg and M Sheves
Center for Biophysics and Computational Biology, University of Illinois at Urbana-Champaign 61801.
Following light absorption, at neutral pH the bacteriorhodopsin mutant Y57N
does not show Schiff base deprotonation (no M intermediate) or proton
pumping activity. We reasoned that this might be due to improper delta pKa
between the proton-donating Schiff base and the proton- accepting Asp-85
after light absorption. To test this, we reduced the intrinsic pKa of the
protonated Schiff base in the pigment (and thus in the photointermediates)
by replacing the retinal chromophore with an analogue, 14-F retinal. This
substitution restores light-induced M formation, strongly suggesting that
light-induced Schiff base deprotonation is accomplished by lowering its pKa
during the photochemical cycle. Thus, while it is generally accepted that
the Schiff base deprotonation during the photocycle takes place because of
the light-induced reduction in its pKa, we provide here the first
experimental evidence of this phenomenon.
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