Biophysics and Physicobiology
Online ISSN : 2189-4779
ISSN-L : 2189-4779
Regular Article
Role of Thr82 for the unique photochemistry of TAT rhodopsin
Teppei SugimotoKota KatayamaHideki Kandori
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2021 Volume 18 Pages 108-115

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Abstract

Marine bacterial TAT rhodopsin possesses the pKa of the retinal Schiff base, the chromophore, at neutral pH, and photoexcitation of the visible protonated state forms the isomerized 13-cis state, but reverts to the original state within 10–5 sec. To understand the origin of these unique molecular properties of TAT rhodopsin, we mutated Thr82 into Asp, because many microbial rhodopsins contain Asp at the corresponding position as the Schiff base counterion. A pH titration study revealed that the pKa of the Schiff base increased considerably in T82D (>10.5), and that the pKa of the counterion, which is likely to be D82, is 8.1. It was thus concluded that T82 is the origin of the neutral pKa of the Schiff base in TAT rhodopsin. The photocycle of T82D TAT rhodopsin exhibited strong pH dependence. When pH is lower than the pKa of the counterion (pH <8.1), formation of the primary K intermediate was observed by low-temperature UV-visible spectroscopy, but flash photolysis failed to monitor photointermdiates at >10–5 sec. The results were identical for the wild-type TAT rhodopsin. In contrast, when pH was higher than the pKa of the counterion, we observed the formation of the M intermediate, which decayed with the time constants of 3.75 ms and 12.2 sec. It is likely that the protonation state of D82 dramatically switches the photoreaction dynamics of T82D, whose duration lies between <10–5 sec and >10 sec. It was thus concluded that T82 is one of the determinants of the unique photochemistry of TAT rhodopsin.

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