Molecular contacts for chlorosome envelope proteins revealed by cross-linking studies with chlorosomes from Chlorobium tepidum
Molecular contacts for chlorosome envelope proteins revealed by cross-linking studies with chlorosomes from Chlorobium tepidum
复制标题
DOI：
10.1021/bi060776y
复制
发表时间：
2006-08-01
期刊：
影响因子：
2.9
通讯作者：
Bryant, Donald A.
中科院分区：
文献类型：
--
作者：
Li, Hui;Frigaard, Niels-Ulrik;Bryant, Donald A.
绿体是一种独特的捕光触角，存在于绿色细菌的两个门：绿藻和绿藻中。在绿色硫磺细菌Chlorobium tepidum中，叶绿体被膜存在10种蛋白质(CSMA、CSMB、CSMC、CSMD、CSME、CSMF、CsmH、CSMI、CsmJ和CsmX)。用零长交联剂1-乙基-3-[3-(二甲氨基)丙基]碳二亚胺(EDC)对野生型和突变体的绿小体进行了交联和凝胶电泳法分析。改变时间和温度，或用戊二醛取代EDC，均可观察到相似的交联产物。用抗重组绿小体蛋白的多克隆抗体进行免疫印迹，鉴定了交联产物中绿小体蛋白之间的特异性相互作用。我们通过证明这些产物在适当的突变体中缺失来证实这些相互作用。确定了CSMA在准晶基板中的位置，交联法表明CSMA形成了十二聚体大小的二聚体、三聚体和同源多聚体，CSMA直接与Fenna-Matthews-Olson蛋白相互作用。交联性进一步表明CSMA的前体形式插入基板边缘附近，CSMA和前CSMA与CSMB和CSMF相互作用。几种绿小体蛋白，包括CSMA、CSMC、CSMD、CsmH、CSMI、CsmJ和CsmX，都以同源多聚体的形式存在于绿小体被膜中。根据从这些交联实验中获得的结构信息，提出了一个关于绿体被膜蛋白质的位置和相互作用的模型。
Chlorosomes are unique light-harvesting antennae found in two phyla of green bacteria: Chlorobi and Chloroflexi. In the green sulfur bacterium Chlorobium tepidum, 10 proteins ( CsmA, CsmB, CsmC, CsmD, CsmE, CsmF, CsmH, CsmI, CsmJ, and CsmX) exist in the chlorosome envelope. Chlorosomes from the wild type and mutants lacking a single chlorosome protein were cross-linked with the zero-length cross-linker 1-ethyl-3-[3-(dimethylamino) propyl] carbodiimide (EDC) and analyzed by gel electrophoresis. Similar cross-linking products were observed when the time and temperature were varied or when EDC was replaced with glutaraldehyde. Specific interactions between chlorosome proteins in cross-linked products were identified by immunoblotting with polyclonal antibodies raised against recombinant chlorosome proteins. We confirmed these interactions by demonstrating that these products were missing in appropriate mutants. Confirming the location of CsmA in the paracrystalline baseplate, cross-linking showed that CsmA forms dimers, trimers, and homomultimers as large as dodecamers and that CsmA directly interacts with the Fenna-Matthews-Olson protein. Cross-linking further suggests that the precursor form of CsmA is inserted near the edges of the baseplate, where CsmA and pre-CsmA interact with CsmB and CsmF. Several chlorosome proteins, including CsmA, CsmC, CsmD, CsmH, CsmI, CsmJ, and CsmX, were shown to exist as homomultimers in the chlorosome envelope. On the basis of the structural information obtained from these cross-linking experiments, a model for the locations and interactions of the proteins of the chlorosome envelope is proposed.