Structure and mechanism in active transport systems for sugars, amino acids and ions.

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Transport of sugars, amino acids and ions across cell membranes is very important for absorption of nutrients, maintenance of intracellular homeostasis and energy transduction in cells. In this project, we tried to analyze Na^+-coupled or H^+-coupled active transport of sugars and amino acids, ATP synthesis, and Na^+/H^+ antiporter from biochemical and genetic aspects.The melibiose transport system of Escherichia coli is an Na^+-coupled system. We isolated many mutants which showed altered cation specificly of the transport carrier. We cloned the mutated gene (melB), and determined the nucleotide sequences. We found that substitutions of some amino acid residues in the melibiose carrier caused alteration in cation specificly. We also analyzed the membrane topology of the melibiose carrier by using phoA-melB fusion, and determined the topology of the carrier in membrane. We also analyzed the Na^+/H^+ antiporter, and found that it was important for intracellular pH regulation. We found Na^+-coupled transport system for branched-chain amino acids in Pseudomonas aeruginosa, and characterized it. We also found an H^+-coupled citrate transport system in Salmonella typhimurium. The gene (citA) was cloned and the nucleotide sequences were determined. Thus, structural characteristics of the H^+-citrate carrier were revealed. Structure and function of an H^+-coupled and an Na^+- coupled ATP synthesizing systems were also analyzed.