Titration kinetics of Asp‐85 in bacteriorhodopsin: exclusion of the retinal pocket as the color‐controlling cation binding site

FEBS Letters

Ottolenghi, M¹  Friedman, N²  Eliash, T²  Fu, X¹  Bressler, S¹  Sheves, M² 

[1]Department of Physical Chemistry, The Hebrew University, Jerusalem 91904, Israel

[2]Department of Organic Chemistry, The Weizmann Institute of Science, Rehovot 76100, Israel

关键词: Purple and blue bacteriorhodopsin;    Titration kinetics of Asp-85;    Metal cation binding site;    bR;    bacteriorhodopsin;    OP;    1;    10-phenanthroline;    bP;    bathophenanthroline disulfonic acid;    Asp;    aspartic acid;    Glu;    glutamic acid;    Et;    ethyl;    Pr;    propyl;    Bu;    butyl;   

DOI  :  10.1016/S0014-5793(97)01194-0

学科分类：生物化学/生物物理

来源: John Wiley & Sons Ltd.

【 摘 要 】

The spectrum (the purple↔blue transition) and function of the light-driven proton pump bacteriorhodopsin are determined by the state of protonation of the Asp-85 residue located in the vicinity of the retinal chromophore. The titration of Asp-85 is controlled by the binding/unbinding of one or two divalent metal cations (Ca²⁺ or Mg²⁺). The location of such metal binding site(s) is approached by studying the kinetics of the cation-induced titration of Asp-85 using metal ions and large molecular cations, such as quaternary ammonium ions, R₄N⁺ (R=Et, Pr, a divalent ‘bolaform ion’ [Et₃N⁺-(CH₂)₄-N⁺Et₃] and the 1:3 molecular complex formed between Fe²⁺ and 1,10-phenanthroline (OP). The basic multi-component kinetic features of the titration, extending from 10⁻² to 10⁴ s, are unaffected by the charge and size of the cation. This indicates that cation binding to bR triggers the blue→purple titration in a fast step, which is not rate-determining. In view of the size of the cations involved, these observations indicate that the cation binding site is in an exposed location on, or close to, the membrane surface. This excludes previous models, which placed the color-controlling Ca²⁺ ion in the retinal binding pocket.

【 授权许可】

Unknown