title: Structure of the N intermediate of bacteriorhodopsin revealed by x-ray diffraction.
abstract: X-ray diffraction experiments revealed the structure of the N photointermediate of bacteriorhodopsin. Since the retinal Schiff base is reprotonated from Asp-96 during the M to N transition in the photocycle, and Asp-96 is reprotonated during the lifetime of the N intermediate, or immediately after, N is a key intermediate for understanding the light-driven proton pump. The N intermediate accumulates in large amounts during continuous illumination of the F171C mutant at pH 7 and 5 degrees Celsius. Small but significant changes of the structure were detected in the x-ray diffraction profile under these conditions. The changes were reversible and reproducible. The difference Fourier map indicates that the major change occurs near helix F. The observed diffraction changes between N and the original state were essentially identical to the diffraction changes reported for the M intermediate of the D96N mutant of bacteriorhodopsin. Thus, we find that the protein conformations of the M and N intermediates of the photocycle are essentially the same, in spite of the fact that in M the Schiff base is unprotonated and in N it is protonated. The observed structural change near helix F will increase access of the Schiff base and Asp-96 to the cytoplasmic surface and facilitate the proton transfer events that begin with the decay of the M state.
authors.affiliation: Department of Physics, Osaka University, Japan.
authors.forename: H
authors.initials: H
authors.lastname: Kamikubo
authors.forename: M
authors.initials: M
authors.lastname: Kataoka
authors.forename: G
authors.initials: G
authors.lastname: Váró
authors.forename: T
authors.initials: T
authors.lastname: Oka
authors.forename: F
authors.initials: F
authors.lastname: Tokunaga
authors.forename: R
authors.initials: R
authors.lastname: Needleman
authors.forename: J K
authors.initials: JK
authors.lastname: Lanyi
mesh_terms: D001224:Aspartic Acid / Q000737:chemistry; D001436:Bacteriorhodopsins / Q000737:chemistry* / Q000235:genetics / Q000528:radiation effects; D005583:Fourier Analysis; D006217:Halobacterium / Q000737:chemistry* / Q000235:genetics; D008565:Membrane Proteins / Q000737:chemistry* / Q000235:genetics / Q000528:radiation effects; D016297:Mutagenesis, Site-Directed; D010777:Photochemistry; D011487:Protein Conformation / Q000528:radiation effects*; D011522:Protons; D011994:Recombinant Proteins / Q000737:chemistry / Q000528:radiation effects; D012545:Schiff Bases / Q000737:chemistry / Q000528:radiation effects; D014961:X-Ray Diffraction
chemical_list: D008565:Membrane Proteins; D011522:Protons; D011994:Recombinant Proteins; D012545:Schiff Bases; D001224:Aspartic Acid; D001436:Bacteriorhodopsins