Chloride Concentration Dependency of the Electrogenic Activity of Halorhodopsin

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• 作者：Daichi Okuno ; Makoto Asaumi ; and Eiro Muneyuki
• 刊名：Biochemistry
• 出版年：1999
• 出版时间：April 27, 1999
• 年：1999
• 卷：38
• 期：17
• 页码：5422 - 5429
• 全文大小：128K
• 年卷期：v.38,no.17(April 27, 1999)
• ISSN：1520-4995

文摘

The capacitive photoelectric current responses of the halorhodopsins from Halobacteriumsalinarum (shR) and from Natronobacterium pharaonis (phR) were studied using membrane fragmentsadsorbed onto a thin polyester film. The electric current of shR was not much affected by ionic strengthor cations present in the medium (Na⁺, K⁺, Li⁺, Mg²⁺, or Ca²⁺), but was greatly influenced by the Cl^-concentration. It increased biphasically as the Cl^- concentration increased from 0 to 5 M, then decreasedand almost vanished at around 10 or 12 M. Apparent K_d's of about 0.1 and 6 M were deduced for the K_dof Cl^- uptake sites. We had to assume a sigmoidal increase of Cl^- binding with a Hill coefficient ofabout 8 at the cytoplasmic, Cl^- release site(s). The half-maximum Cl^- concentration for the sigmoidalbinding was about 7.5 M. The electric current of phR had a maximum around 30 mM Cl^- and biphasicallydecreased at higher Cl^- concentrations. The apparent K_d for the Cl^- uptake site was 5 mM. The biphasicdecrease in the transport activity was explained by assuming a sum of simple hyperbolic type binding (K_d= 0.2 M) and sigmoidally increasing binding with a Hill coefficient of 10 on the cytoplasmic side. Thehalf-maximum concentration of the latter cooperative binding was 5.6 M. This great difference betweenthe apparent affinity of the release site of shR and that of phR can explain the previously reported differencebetween the Cl^- dependency of their photocycles. These results also suggest that there may be multipleCl^- binding sites in the Cl^- transport pathway. A simple sequence of Cl^- transport steps based on amultiion channel model is proposed.