Abstract
Bacteriorhodopsin (bR), a membrane protein found in Halobacterium salinarum, contains retinal as chromophore and functions as a light-driven proton pump. We have carried out four-dimensional X-ray crystallographic studies of bR and its homologous proteins, to elucidate the proton pumping mechanism. A novel crystallization method, called the membrane fusion method, has been developed to prepare a 3D crystal that is stable over a wide pH range. This enabled us to investigate pH-induced as well as light-induced structural changes. Structural analyses of reaction intermediates revealed that water relocation, which affects the pKa values of ionizable residues in the proton channel, takes place in the early stage of the proton pumping cycle. On the basis of this observation, we hypothesized that bR is a water/proton anti-porter. Since some of internal cavities are highly conserved during the evolution, it is suggested that their morphological changes initiated by the retinal photo-isomerization play an important role for water relocation and proton tranlocation across the membrane.
