STABLE PHOTOBLEACHING OF P840 IN CHLOROBIUM REACTION-CENTER PREPARATIONS - PRESENCE OF THE 42-KDA BACTERIOCHLOROPHYLL ALPHA-PROTEIN AND A 17-KDA POLYPEPTIDE
STABLE PHOTOBLEACHING OF P840 IN CHLOROBIUM REACTION-CENTER PREPARATIONS - PRESENCE OF THE 42-KDA BACTERIOCHLOROPHYLL ALPHA-PROTEIN AND A 17-KDA POLYPEPTIDE
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DOI：
10.1021/bi00029a039
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发表时间：
1995-07-25
期刊：
影响因子：
2.9
通讯作者：
NELSON, N
中科院分区：
文献类型：
--
作者：
HAGERBRAUN, C;XIE, DL;NELSON, N
本文提出并定量了以赤潮氯杆菌和利氏氯杆菌为原料，高产率厌氧制备光活性稳定的P840反应中心的简便方法。蔗糖密度梯度共沉淀法测定亚基组成。青霉的P840反应中心蛋白PSCA、BChla天线蛋白FMO、中心A和中心B的FeS蛋白PSCB和带正电的17 kDa蛋白PSCD是最小的四个亚基，从毛白菌制备的PSCC还含有细胞色素c-551。纯化后的配合物的BChla吸收峰位于810 nm，肩峰位于835 nm。如果假定BChla的消光系数是均匀的，则肩峰比为0.25，相当于每70BChla分子有一个反应中心。然而，在610 nm的连续光下漂白相当于每50个BChla分子对应一个光活性反应中心。因此，要么高估了BChla在反应中心的消光系数，要么低估了光漂白的消光系数。在任何情况下，反应中心的主要部分在准备过程中是光活性的。副产物为P840反应中心亚复合体，缺失PSCD，缺失FMO和PSCB，但保留细胞色素c亚基。它是光敏性的，在814 nm处有一个吸收峰，835/814的吸光度比为0.42。FMO和PSCB有形成互补亚复合体的趋势。FMO和PSCD显然是稳定光活性反应中心所必需的，而细胞色素c亚单位则不是。
Simple procedures for the anaerobic preparation of photoactive and stable P840 reaction centers from Chlorobium tepidum and Chlorobium limicola in good yield are presented and quantitated. The subunit composition was tested by cosedimentation in sucrose density gradients. For C. limicola, it minimally comprises four subunits: the P840 reaction center protein PscA, the BChla antenna protein FMO, the FeS protein PscB with centers A and B, and a positively charged 17-kDa protein denoted PscD, The preparation from Chlorobium tepidum additionally contained PscC, a cytochrome c-551. The BChla absorption peak of the purified complexes was at 810 nm, with a shoulder at 835 nm. The ratio of the shoulder to the peak was 0.25, which corresponds to 1 reaction center per 70 BChla molecules if a uniform extinction coefficient of BChla is assumed. However, bleaching at 610 nm in continuous light corresponded up to 1 photoactive reaction center per 50 BChla molecules. Therefore, either the extinction coefficient of BChla in the reaction center is overestimated or the one for photobleaching is underestimated. In any case, the major portion of the reaction center was photoactive in the preparations. A P840 reaction center subcomplex, lacking PscD and deficient in FMO and PscB, but retaining the cytochrome c subunit, was obtained as a side product. It was photoinactive and had an absorption peak at 814 nm and a 835/814 absorbance ratio of 0.42. FMO and PscB show the tendency to form a complementary subcomplex. FMO and PscD are apparently required to stabilize the photoactive reaction center, while the cytochrome c subunit is not.