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Effect of Substitution of Proline-77 to Aspartate on the Light-Driven Proton Release of Bacteriorhodopsin

Authors

  • Yazhuo Wang,

    1. State Key Laboratory of Molecular Engineering of Polymers, Department of Macromolecular Science, Advanced Materials Laboratory, Fudan University, Shanghai, China
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  • Yingchun Zhao,

    1. State Key Laboratory of Molecular Engineering of Polymers, Department of Macromolecular Science, Advanced Materials Laboratory, Fudan University, Shanghai, China
    2. Department of Biochemistry, School of Life Sciences, Fudan University, Shanghai, China
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  • Ming Ming,

    1. State Key Laboratory of Molecular Engineering of Polymers, Department of Macromolecular Science, Advanced Materials Laboratory, Fudan University, Shanghai, China
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  • Jia Wu,

    1. State Key Laboratory of Molecular Engineering of Polymers, Department of Macromolecular Science, Advanced Materials Laboratory, Fudan University, Shanghai, China
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  • Weida Huang,

    1. Department of Biochemistry, School of Life Sciences, Fudan University, Shanghai, China
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  • Jiandong Ding

    Corresponding author
    1. State Key Laboratory of Molecular Engineering of Polymers, Department of Macromolecular Science, Advanced Materials Laboratory, Fudan University, Shanghai, China
      Corresponding author email: jdding1@fudan.edu.cn (Jiandong Ding)
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Corresponding author email: jdding1@fudan.edu.cn (Jiandong Ding)

Abstract

Wild-type bacteriorhodopsin (BR) and another retinal protein archaerhodopsin 4 (AR4) are both light-driven proton pumps, but exhibit opposite temporal orders of proton release and uptake upon a flash illumination at neutral pH due to a higher pKa of proton release complex (PRC) in AR4. Since the 77th residue in the extracellular side is proline (P) in BR, but aspartic acid (D) in AR4, we have mutated P77 in BR by D in this study. The new point mutation was found to affect the kinetics of proton release and the pH dependence significantly. Upon a flash excitation, three components “fast proton release,”“proton uptake” and “slow proton release” were observed at neutral pH in P77D. The pKa of PRC in the M intermediate was increased from 5.6 in the wild-type to 7.0, and became closer to that in AR4, which is 8.4. The coupling strength between D85 and PRC were also weakened, as expected. These data indicate that the 77th residue in AR4 greatly account for the difference between the two proton pumps.

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