Structure of the N intermediate of bacteriorhodopsin revealed by x-ray diffraction

Hironari Kamikubo, Mikio Kataoka, György Váró, Toshihiko Oka, Fumio Tokunaga, Richard Needleman, Janos K. Lanyi

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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°C. 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.

Original languageEnglish
Pages (from-to)1386-1390
Number of pages5
JournalProceedings of the National Academy of Sciences of the United States of America
Issue number4
Publication statusPublished - Feb 20 1996

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