Crystal Structure of the 11-cis Isomer of Pharaonis Halorhodopsin: Structural Constraints on Interconversions among Different Isomeric States
详细信息 查看全文
- 作者：Siu Kit Chan ; Haruki Kawaguchi ; Hiroki Kubo ; Midori Murakami ; Kunio Ihara ; Kosuke Maki ; Tsutomu Kouyama
- 刊名：Biochemistry
- 出版年：2016
- 出版时间：July 26, 2016
- 年：2016
- 卷：55
- 期：29
- 页码：4092-4104
- 全文大小：684K
- 年卷期：0
- ISSN：1520-4995
文摘
Like other microbial rhodopsins, the light driven chloride pump halorhodopsin from Natronomonas pharaonis (pHR) contains a mixture of all-trans/15-anti and 13-cis/15-syn isomers in the dark adapted state. A recent crystallographic study of the reaction states of pHR has shown that reaction states with 13-cis/15-syn retinal occur in the anion pumping cycle that is initiated by excitation of the all-trans isomer. In this study, we investigated interconversions among different isomeric states of pHR in the absence of chloride ions. The illumination of chloride free pHR with red light caused a large blue shift in the absorption maximum of the retinal visible band. During this “red adaptation”, the content of the 11-cis isomer increased significantly, while the molar ratio of the 13-cis isomer to the all-trans isomer remained unchanged. The results suggest that the thermally activated interconversion between the 13-cis and the all-trans isomers is very rapid. Diffraction data from red adapted crystals showed that accommodation of the retinal chromophore with the 11-cis/15-syn configuration was achieved without a large change in the retinal binding pocket. The measurement of absorption kinetics under illumination showed that the 11-cis isomer, with a λmax at 565 nm, was generated upon excitation of a red-shifted species (λmax = 625 nm) that was present as a minor component in the dark adapted state. It is possible that this red-shifted species mimics an O-like reaction state with 13-cis/15-syn retinal, which was hypothesized to occur at a late stage of the anion pumping cycle.