RUI: The Use of Antibodies to Study the Topology of Thylakoid Protein Complexes

从菠菜类囊体膜中分离出三种蛋白复合物表面决定因子的多克隆和单克隆抗体，通过对正侧边或内侧边类囊体膜的亲和吸附分离得到。这些侧特异性抗体将用于识别那些在类囊体悬浮的水环境中显示抗原决定因子的蛋白质。比较基质暴露的蛋白质和管腔暴露的蛋白质将表明哪些蛋白质是跨膜的。在其他实验中，单克隆抗体将用于证明哪些蛋白质位于与水氧化有关的四个腔侧外源蛋白质之下。总之，这些实验将极大地增强我们对所选蛋白质复合物拓扑结构的理解，从而允许开发越来越复杂的光合作用能量守恒过程的结构/功能模型。这些实验的总体目标是增加我们对光合能量耦合所需的三种蛋白质复合物的横向取向和拓扑结构的理解。提出的研究将促进这些蛋白质复合物的更复杂和拓扑正确模型的发展，并导致对光合作用的更好理解。

Polyclonal and monoclonal antibodies specific for surface determinants of three protein complexes isolated from spinach thylakoid membranes will be isolated by affinity adsorption to rightsideout or insideout thylakoid membranes. These side- specific antibodies will be used to identify those proteins which display antigenic determinants to the aqueous milieu in which the thylakoids are suspended. Comparison of those proteins with stromal exposures with those proteins displaying lumen exposures will indicate which proteins are transmembranous. In other experiments, monoclonal antibodies will be used to demonstrate which proteins lie beneath the four lumen side extrinsic proteins which are involved with water oxidation. Together, these experiments will greatly enhance our understanding of the topology of the selected protein complexes, thus allowing the development of increasingly sophisticated structure/function models of the process of photosynthetic energy conservation. %%% The overall objective of these experiments is to increase our understanding of the transverse orientations and topologies of three protein complexes required for photosynthetic energy coupling. The proposed research will enhance the development of more sophisticated and topologically correct models of these protein complexes and lead to a better understanding of photosynthesis.