Membrane Transport of Melibiose by Escherichia coli

本研究的目的是阐明载体分子在细菌中底物-阳离子共转运系统中的作用机制，特别是对黑二糖载体的研究。分离缺乏阳离子识别和糖识别的自发突变体之后，将分析它们在膜运输中的生理异常。我们将研究载体分子中氨基酸取代的生理效应，特别是在跨膜区域。通过使用定点突变技术，有望对“螺旋发夹”假说进行实验检验，该假说是将黑二糖载体插入细胞膜。其他研究将使用多克隆抗体，试图确定假定的载体在膜上的“环”区域的方向。糖和无机离子等小分子通过富含脂质的细胞膜进入细胞的方式是一个基本的感兴趣和重要的问题；跨膜的主动运输通常涉及特定的载体分子，而它就是其中之一--二糖载体--这是本研究的重点。该项目的目标是了解载体分子的分子方面，使其能够特定地与某些分子结合，而不是其他分子，并以允许糖和阳离子沿适当方向共运输的方式插入细胞膜。//

The goal of the research is to elucidate the mechanism of action of the carrier molecule in a substrate-cation cotransport system in the bacterium E. coli, with specific focus on the melobiose carrier. Isolation of spontaneous mutants deficient in cation recognition and sugar recognition will be followed by analysis of their physiological abnormalities in membrane transport. The physiological effects of amino-acid substitution in the carrier molecule, particularly in membrane-spanning regions, will be examined. Use of site-directed mutagenesis is expected to provide an experimental test of the "helical hairpin" hypothesis for insertion of the melobiose carrier into membranes. Other studies will employ polyclonal antibodies in an attempt to determine the orientation of presumed "loop" regions of the carrier in the membrane. The means whereby small molecules such as sugars and inorganic ions gain entrance to cells through the lipid-rich cell membrane is a matter of fundamental interest and importance; active transport across membranes often involves specific carrier molecules and it is one of these--the melobiose carrier--that is the focus of this research. The goal of the project is to understand the molecular aspects of the carrier molecule that permit it to bind specifically to some molecules and not others, and to be inserted into cell membranes in a manner that permits cotransport of sugars and cations in appropriate directions. //