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

放氧膜制剂含有多种内源多肽，包括约47 kDa和43-40 kDa的两个携带叶绿素的亚基多肽，33、24和18 kDa的三个参与水氧化的外源蛋白，以及高等植物中捕光的叶绿素a/b蛋白和蓝藻中的藻胆蛋白等天线色素复合体。在这项研究中，我们通过可逆交联法检测了两种放氧制剂的亚基多肽之间的最近邻关系。以可裂解的双官能团交联剂丁二酰丙酸酯为交联剂，研究了菠菜放氧膜亚单位多肽之间的最近邻关系。在与水氧化有关的三种外源蛋白中，33 kDa的蛋白与光系统II反应中心复合体的47 kDa亚基多肽发生交联，而18 kDa和24 kDa的蛋白相互交联。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。还考察了聚球藻放氧颗粒中保留少量别藻蓝蛋白的多肽之间的最近邻关系。交联产物的二硫键的交联和随后的断裂表明，在77K发射长波荧光带的别藻蓝蛋白物种与颗粒交联。双向琼脂糖凝胶电泳显示，藻胆蛋白与光系统II反应中心复合体中携带40 kDa叶绿素的蛋白发生了特异性的交联。结果表明，40 kDa的脱脂蛋白在藻胆体向光系统II反应中心的激发能量转移中起着关键作用。

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.