Water molecule rearrangements around Leu93 and Trp182 in the formation of the L intermediate in bacteriorhodopsin's photocycle

Akio Maeda, Farol L. Tomson, Robert B. Gennis, Sergei P. Balashov, Thomas G. Ebrey

Research output: Contribution to journalArticlepeer-review


After the chromophore's isomerization in the initial photochemical event in bacteriorhodopsin, the primary photoproduct K makes a thermal transition to the L intermediate, which prepares the pigment for Schiff base deprotonation in the following step (L → M). Substantial changes in the hydrogen bonding of internal water molecules take place upon L formation. Some of these mobile waters are probably involved in changing the pK of the Schiff base and perhaps that of the proton acceptor Asp85 to allow proton movement [Maeda, A. (2001) Biochemistry (Moscow) 66, 1555-1569]. Here we show that mutations of Leu93 and Trp182, residues close to the 13-methyl group of the chromophore, allow the formation of L at much lower temperatures than in the wild type (80 K instead of 140 K). Moreover, an intense band due to weakly bound water that is peculiar for L was already present in the initial (unphotolyzed) state of each mutant at 2632 cm-1 (in D2O) but not in the wild type. This unique, intense water band is shifted compared to the L band at 2589 cm-1 but coincides with the band seen in L', the all-trans photoproduct of wild-type L formed at 80 K. We propose that the L93M and W182F mutations induce changes in the hydrogen bonding of one or more water molecules in the unphotolyzed states of these pigments, which are similar to those H-bonding changes that take place upon formation of L in the wild type, and thus facilitate the formation of L even at 80 K. We infer that L formation involves perturbation of a site which includes retinal, Trp182, and Leu93, and this structure is temporarily stabilized by rearranged hydrogen bonds with water molecules.

Original languageEnglish (US)
Pages (from-to)2535-2541
Number of pages7
Issue number9
StatePublished - Mar 11 2003

ASJC Scopus subject areas

  • Biochemistry


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