Ariki M, Lanyi J K
J Biol Chem. 1986 Jun 25;261(18):8167-74.
We have investigated the effects of the binding of various metal ions to cation-free bacteriorhodopsin ("blue membrane"). The following have been measured: shift of the absorption maximum from 603 to 558 nm (blue to purple transition), binding isotherms, the release of H+ upon binding, and the decay of the deprotonated intermediate of the photocycle, M412. We find that all cations of the lanthanide series, as well as the alkali and alkali earth metals earlier investigated, are able to bring about the absorption shift, whereas Hg2+ and Pt4+ are not. Sigmoidal spectroscopic titration curves and nonsigmoidal binding curves suggest that there are two high affinity sites for cations in bacteriorhodopsin. Binding to the site with the second highest affinity is responsible for the absorption shift. Divalent cation binding to blue membrane causes release of about six protons, whereas higher numbers of protons are released by trivalent cations, suggesting that the shift of absorption maximum involves proton release from carboxyl group(s). The metal ion bound to this site must be surrounded by carboxyl oxygen atoms acting together as a multidentate ligand with a specific geometry because multivalent ions are effective only when capable of octahedral coordination. Lanthanide ions dramatically inhibit M412 decay at pH above 6.3, an effect probably due to binding to lipid phosphoryl groups.
我们研究了各种金属离子与无阳离子细菌视紫红质（“蓝膜”）结合的影响。已测量了以下各项：吸收峰最大值从603nm移至558nm（从蓝色到紫色转变）、结合等温线、结合时H⁺的释放以及光循环中去质子化中间体M412的衰减。我们发现镧系元素的所有阳离子，以及先前研究过的碱金属和碱土金属，都能够引起吸收峰的移动，而Hg²⁺和Pt⁴⁺则不能。S形光谱滴定曲线和非S形结合曲线表明，细菌视紫红质中存在两个阳离子高亲和力位点。与第二高亲和力位点的结合导致了吸收峰的移动。二价阳离子与蓝膜的结合会导致约六个质子的释放，而三价阳离子释放的质子数更多，这表明吸收峰最大值的移动涉及羧基的质子释放。结合到该位点的金属离子必须被羧基氧原子包围，这些羧基氧原子共同作为具有特定几何形状的多齿配体起作用，因为多价离子只有在能够进行八面体配位时才有效。镧系离子在pH高于6.3时显著抑制M412的衰减，这种效应可能是由于与脂质磷酸基团的结合。
