Deformation of Helix C in the Low Temperature L-intermediate of Bacteriorhodopsin

Abstract

Light-driven vectorial proton translocation is basic to the mechanism of energy transduction by photosynthetic systems. Bacteriorhodopsin (bR)1 is the simplest known light-driven proton pump and has long served as a model system for understanding how protons may be transported "up hill" against a transmembrane proton motive potential. bR contains seven transmembrane ␣-helices that surround a proton translocation channel lined with strategically placed charged residues (3). Depending upon their protonation states, which change in a well orchestrated cascade as a proton is transported across the cell membrane, these charged residues can serve as either proton donors or proton acceptors. Light activation of the chromophore, an all-trans-retinal molecule covalently attached to Lys-216 in helix G via a protonated Schiff base (the primary proton donor) results in the 13-cis-retinal configuration with two-thirds quantum efficiency. Steric conflicts and mechanical stress resulting from photoisomerization initiate a sequence of conformational changes that can be characterized spectroscopically and that perturb the local environment of several key residues, strongly affecting their pK a values and creating transient pathways for proton transfer.The specific spectral intermediates of the bR photocycle have been well characterized, and a common reaction scheme is: bR 570 3 K 590 7 L 550 7 M 412 7 N 560 7 O 640 3 bR 570 (sub-

Mechanism of a proton pump analyzed with computer simulations

“…The millisecond timescale associated with the long-distance proton transfers may be required for protein conformational changes. The crystal structures of the reaction cycle intermediates indicate rather small structural rearrangements of the protein in K [3,4], L [5][6][7][8][9], and early-M [10]; there are structural rearrangements of the protein during the M-to-N [11][12][13][14][15], and N-to-O transitions [16][17][18]. The passage of the protein through the intermediate conformational states is also accompanied by changes in the number and location of internal water molecules [19,20].…”

“…A cytoplasmic orientation of the retinal prior to deprotonation is supported by the crystal structures of the L-state models from Refs. [5][6][7] (see iceblue and purple structures in Fig. 3a), FTIR experiments [19], and by the good agreement between the bR-L dipole moment change obtained from experiments and that computed with the L-state model structure from Ref.…”

“…The importance of the bridging water molecule is suggested by the observation that in its absence direct hydrogen bonds between the Schiff base and D85 form only transiently ( [6]; Bondar and Smith, unpublished results). Moreover, the persistence of a hydrogen bond between the Schiff base and wB after proton transfer [76] is consistent with the observation from NMR that in early M the deprotonated Schiff base hydrogen bonds with a hydroxyl group [65].…”

“…3b). Although retinal is a flexible molecule and it samples a relatively wide range of conformations during dynamics [6,38,71,72], it is unclear if all geometries proposed in the L-state crystal structures [5][6][7][8][9] are indeed sampled during the reaction path. The significant energy barrier for retinal isomerisation from 15-trans to 15-syn [32,46] makes it unlikely that a 13-cis, 15-syn geometry [8] could be reached during the K-to-L transition [33].…”

“…a, b comparison of L-state X-ray structural models with the bR resting state. The following color codes are used: green bR resting state [1,103]; iceblue 1VJM [6]; magenta 1UCQ [7]; blue 2NTW [9]; mauve 1O0A [8]. The Schiff base nitrogen atom is shown as a sphere.…”