Topographical studies on subunit polypeptides of oxygen-evolving PS II preparations by reversible crosslinking.

进化氧膜含有多种内在多肽，包括2种携带叶绿素的亚基多肽，长度分别为47和43-40 kDa， 3种外源蛋白，长度分别为33、24和18 kDa，涉及水氧化和天线色素复合物，如高等植物中的光收集叶绿素a/b蛋白（LHCP）和蓝藻中的藻胆蛋白。在这项研究中，我们通过可逆交联研究了两种出氧制剂的亚基多肽之间的近邻关系。采用可裂解的双功能交联试剂dithiobis（琥珀酰丙酸酯）研究了菠菜进化氧膜亚基多肽之间的近邻关系，最大分子跨度为12 < 1>。在与水氧化相关的三种外源蛋白中，33 kDa蛋白与光系统II反应中心复合物的47 kDa亚基多肽交联，而18和24 kDa蛋白相互交联。33 kDa蛋白与47 kDa蛋白的结合表明后者在水氧化中起重要作用。我们还研究了含少量异藻蓝蛋白的聚藻球菌进化氧颗粒多肽之间的近邻关系。交联产物的交联和随后的二硫键断裂表明，在77K下发出长波长荧光带的异藻蓝蛋白与颗粒交联。二维SDS聚丙烯酰胺凝胶电泳显示，藻胆蛋白与光系统II反应中心复合物中携带叶绿素的40 kDa蛋白特异性交联。结果表明，40 kDa载脂蛋白在从藻胆小体到光系统II反应中心的激发能转移中起关键作用。

Oxygen-evolving membrane preparations contain several intrinsic polypeptides including two chlorophyll-carrying subunit polypeptides of about 47 and 43-40 kDa, three extrinsic proteins of 33, 24 and 18 kDa involving in water oxidation and antenna pigment complexes such as light-harvesting chlorophyll a/b proteins (LHCP) in higher plants and phycobiliproteins in cyanobacteria. In this study, we examined the nearest neighbor relationships among subunit polypeptides of two oxygen-evolving preparations by reversible crosslinking.1. The nearest neighbor relationships among subunit polypeptides of spinach oxygen-evolving membrane were studied with a cleavable bifunctional crosslinking reagent, dithiobis(succinimidyl propionate) with a maximum molecular span of 12 <ang> . Of the three extrinsic proteins which are associated with water oxidation, the 33 kDa protein was found to crosslink with the 47 kDa subunit polypeptide of the photosystem II reaction center complex, whereas the 18 and 24 kDa proteins crosslinked with each other. The association of the 33 kDa protein with the 47 kDa protein suggests that the latter protein plays an important role in water oxidation.2. The nearest neighbor relationships among polypeptides of synechococcus oxygen-evolving particles which retains a small amount of allophycocyanin were also examined. Crosslinking and subsequent cleaving of disulfide bonds of the crosslinked product showed that an allophycocyanin species which emits a long wavelength fluorescence band at 77K is crosslinked with the particles. Two-dimensional SDS polyacrylamide gel electrophoresis revealed that the phycobiliprotein is specifically crosslinked with a chlorophyll-carrying 40 kDa protein of the photosystem II reaction center complex. The results suggest that the 40 kDa apoprotein plays a key role in the excitation energy transfer from phycobilisomes to the photosystem II reaction center.