Abstract :

It will be attempted to isolate and characterize all necessary components of a galactose transport system of Escherichia coli mediated by the periplasmic galactose-binding protein. The major approach will involve the construction of isogenic strains which carry mutations only in the genes necessary for the Beta-methylgalactoside transport system. The proteins of 3 different fractions isolated from the envelope of these mutants will be separated on two dimensional gel electrophoresis. Transport component will be detected by differential isotope labeling using wild-type proteins as reference. The mechanism of the cell division dependent regulation of the transport component's synthesis will be studied. Ribosomes of a mutant temperature sensitive in cell division will be isolted after growth at the permissive and non-permissive temperature. Alterations of any one ribosomal protein will be detected by two dimensional polyacrylamide gel electrophoresis. An in vitro system will be worked out for the synthesis of the galactose-binding protein. Restoration of galactose-binding protein mediated transport in membrane vesicles will be attempted.