Figure 6. Molecular dynamics simulation and prediction of water molecule distribution of dark states and O-intermediates of BR.

(a) The dark state of BR-WT with separated areas of mobile water. (b) In the O-state of BR-WT, the downward (extracellular side) movement of the guanidine side chain of R82 is depicted. The hydrogen bond connecting the guanidine group with Y57 and D212 was interrupted due to this downward movement. (c) Mutation of R82 by the smaller and neutral Q82 induced a large space with an extended water cluster. (d) Extended water cluster was still present in the O-state of R82Q, but direct accessibility to the primary proton donor D85 was interrupted. (e) Mutation of Y57 by the positively charged K57 promoted a salt bridge between D212 and K57. The dark state of K57Y was also characterized by an increased water cluster. (f) The O-state of Y57K lacks an interaction between R82 and E204. The huge cavity of mobile water molecules between K57, R82, D85, and D212 caused accessibility to the Schiff Base. Downward movement of the guanidine side chain of R82 was also observed.