Time-Resolved Fourier Transform Infrared Study of Structural Changes in the Last Steps of the Photocycles of Glu-204 and Leu-93 Mutants of Bacteriorhodopsin

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• 作者：Hideki Kandori ; Yoichi Yamazaki ; Minoru Hatanaka ; Richard Needleman ; Leonid S. Brown ; Hans-Thomas Richter ; Janos K. Lanyi ; and Akio Maeda
• 刊名：Biochemistry
• 出版年：1997
• 出版时间：April 29, 1997
• 年：1997
• 卷：36
• 期：17
• 页码：5134 - 5141
• 全文大小：154K
• 年卷期：v.36,no.17(April 29, 1997)
• ISSN：1520-4995

文摘

The last intermediate in the photocycle of thelight-driven proton pump bacteriorhodopsin isthe red-shifted O state. The structure and dynamics of the laststep in the photocycle were characterizedwith time-resolved Fourier transform infrared spectroscopy of themutants of Glu-204 and Leu-93, whichaccumulate this intermediate in much larger amounts than the wild type.The results show that E204Qand E204D give distorted all-trans-retinal chromophore likethe O intermediate of the wild type. This issimply due to the perturbation of the proton acceptor function ofGlu-204 in the O-to-BR transition in theGlu-204 mutants. The corresponding red-shifted intermediates ofL93M, L93T, and L93S have a 13-cischromophore like the N intermediate of the wild type, as reported fromanalysis of extracted retinal[Delaney, J. K., Schweiger, U., & Subramaniam, S. (1995) Proc.Natl. Acad. Sci. U.S.A. 92, 11120-11124]. In spite of their different chromophore structures fromthe O intermediate, the red-shiftedintermediates are similar to the O intermediate but not to the Nintermediate of the wild type with respectto structural changes in the peptide carbonyls. The structuralchanges around Asp-96 in the N intermediateare completely restored also in the red-shifted intermediates of theLeu-93 mutants like in the O intermediate.These results imply that the protein structural changes in thelast step proceed regardless of thermalisomerization of the chromophore. Time-resolved Fourier transforminfrared spectroscopy with the Glu-204 mutants suggests that the response of Asp-204 (Glu-204 in the wildtype) to the protonation of Asp-85 during formation of the M intermediate, which results in protonrelease, is slow and may occur throughstructural changes.