No abstract

Kinetics were determined for the four transients K590, L540, M410, O660 of the photochemical cycle of bacteriorhodopsin (BR570) both in 1H2O and in 2H2O over a wide temperature range. Breaks in the Arrhenius plots, observed at 25 degrees-32 degrees for the longest-lived transients coincide with a transition point in the microviscosity of the membrane as measured by depolarization of an added fluorescent probe. The earliest isotope effect occurs in the decay of L540, and is present in the subsequent formation and decay of M410 and O660. Thus in the light-driven proton pump of BR570, proton ejection from the Schiff base correlates with decay of L540 and reprotonation occurs with the decay of both M410 and O660 back to BR570.

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A mode‐locked Nd:YAG laser was used to excite the aromatic amino acid residues of bacteriorhodopsin in the purple membrane and the tryptophan (Trp) fluorescence decay analyzed with a streak camera (λ> 380 nm). The decay kinetics are resolvable into two first‐order half‐times (1.5 and 0.17 ns, respectively), while for retinylidene‐free bacterioopsin, only the longer‐lived Trp emission was observed. The shorter‐lived species reappeared upon regeneration of bacteriorhodopsin by addition of retinal to bacterioopsin but not on treatment of the latter with an equivalent of retinol. It is proposed that these results are consistent with a structural model in which the 7‐8 Trp's distributed among sections A, C, E and F of the seven helical segments A‐G of native bacteriorhodopsin are distinguishable by their distances from the chromophore. Assuming a Förster mechanism for energy transfer with Ro= 25 and 32 Å, respectively, for retinylidene chromophore and retinol the Trp's may be divided into two groups: (i) those completely quenched by retinol and partly quenched by retinal (τ= 0.17) with R ≃ 18 Å and (ii) those (τ= 1.5 ns) which are quenched neither by chromophore nor retinol with R > ca. 30 Å. These results are consistent with and support some of the best models of Engelman et al. (1980) for the protein conformation in the purple membrane.

Kinetics have been determined for the decay of the terminal phototransients M(410) and O(660) and the reappearance of the BR(570) chromophore in flash-photolyzed aqueous suspensions of light-adapted purple membrane fragments from Halobacterium halobium. The results were fitted to a linear model of A k2 M(410)-O(660) -