The Reduction and Restoration of Galactose Transport in Osmotically Shocked Cells of Escherichia coli

Abstract d-Galactose transport in two mutant strains of Escherichia coli K12 lacking galactokinase has been described. These cells showed a reduction of about 50% in the capacity for galactose uptake and in the rate of exit as a consequence of osmotic shock. In the shock procedure the cells were first treated with 0.5 m sucrose containing 1 x 10-4 m ethylenediaminetetraacetate, after which the pelleted cells were rapidly dispersed in cold 5 x 10-4 m MgCl2. A factor was released into the MgCl2 solution (shock fluid) which formed a complex with galactose in vitro, detected by Sephadex chromatography. The material was heat labile and nondialyzable and behaved in several respects like a protein. More of this material appeared when cells high in galactose transport activity were used. The reduced uptake of galactose in shocked cells could be restored by first incubating them with dialyzed shock fluid. The shock fluid could be removed before measuring uptake, suggesting that some factor in the shock fluid needed for uptake could reassociate with the shocked cells. Restoration of activity was always limited to an amount less than the original level. These observations suggest that a factor necessary for galactose transport has been partially released by osmotic shock into the shock fluid. Further, the results suggest that the factor can reassociate with the cell, thereby restoring the reduced uptake of galactose nearly to the original level, but not exceeding it.