Absorption spectral properties of purified halorhodopsin.
Absorption spectral properties of purified halorhodopsin.
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纯化的盐视紫红质的吸收光谱特性。
DOI：
10.1093/oxfordjournals.jbchem.a134695
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发表时间：
1984
影响因子：
2.7
通讯作者：
T. Yoshizawa
中科院分区：
文献类型：
--
作者：
T. Ogurusu;A. Maeda;T. Yoshizawa
盐生盐杆菌Y1膜碎片中的卤视紫红质在576 nm处有一吸收带，该吸收带在0℃红光照射下强度降低(Ogurusu，T.，Maeda，A.，Sasaki，N，&Yoshizawa，T.(1981)J.Biochem)。90、1267-1273)。利用这种光漂白特性作为测定卤视紫质的基础，我们用Triton X-100从膜上提取了卤视紫质，然后用辛基-琼脂糖柱层析纯化了卤视紫质。在NaDodSO4-聚丙烯酰胺凝胶电泳法中，Hr为主带，表观分子量为22,000，但制备物仍有其他几条微弱的条带。纯化的卤视紫红质在1M的氯化钠溶液中，在576 nm处有一个主吸收带，在415 nm附近有一个小吸收带。纯化的卤视紫质在0℃和-75℃下的光反应与膜碎片中的卤视紫质相似。在0℃的红光照射下，纯化的卤视紫质在576 nm附近的吸光度降低，而在410 nm附近的吸光度相应增加。卤视紫红质的吸收光谱与氯离子浓度的依赖关系表明，卤视紫红质只有一个氯离子结合位，其占据是修饰光谱的关键。
Halorhodopsin in the membrane fragments of Halobacterium halobium Y1 showed an absorption band at 576 nm, the intensity of which decreased on irradiation with red light at 0 degrees C (Ogurusu, T., Maeda, A., Sasaki, N., & Yoshizawa, T. (1981) J. Biochem. 90, 1267-1273). Using this photobleachable property as the basis for an assay of halorhodopsin, we purified halorhodopsin by octyl-Sepharose column chromatography after extracting it from the membrane with Triton X-100. In NaDodSO4-polyacrylamide gel electrophoresis, hR appeared as a major band with an apparent molecular weight of 22,000, but the preparation still showed several other faint bands. The purified halorhodopsin showed a main absorption band at 576 nm and a small band at around 415 nm in 1 M NaCl. The photoreactions of the purified halorhodopsin at 0 degrees C and at -75 degrees C were similar to those of halorhodopsin in membrane fragments. Irradiation of the purified halorhodopsin with red light at 0 degrees C resulted in a decrease of absorbance at around 576 nm with a concomitant increase of absorbance at around 410 nm. A hypsochromic photoproduct was obtained on irradiation with 650 nm light at -75 degrees C. The dependency of the absorption spectrum of halorhodopsin on the concentration of chloride indicates that halorhodopsin has a single chloride binding site, occupation of which is responsible for modifying the spectrum.