Mechanisms of Active Transport in Isolated Bacterial Membrane Vesicles

The activity of the α-glycerol-phosphate dehydrogenase-coupled amino acid transport systems in membrane vesicles prepared from Staphylococcus aureus is comparable to the transport activity of intact cells. Temperature optima for initial velocities and steady state levels of amino acid accumulation in the vesicles are at approximately 45° and 25°, respectively, and apparent Michaelis constants for the transport of 17 amino acids are in the micromolar range. There are 12 distinct and specific transport systems for the following families of structurally related amino acids: alanine and glycine; leucine, isoleucine, and valine; serine and threonine; aspartate and glutamate; asparagine and glutamine; lysine; histidine; arginine; phenylalanine, tyrosine and tryptophan; cysteine; methionine; and proline. Structurally dissimilar amino acids do not alter the kinetic constants for the uptake of a given amino acid, and Ki values for amino acids of the same family are essentially identical with the Michaelis constants for the transport of those amino acids. These findings indicate that structurally related amino acids of the same group are transported by the same membrane carrier. Except for the alanine-glycine, leucine-isoleucinevaline, and aspartate-glutamate transport systems in which d-amino acids inhibit the uptake of the appropriate 14C-labeled l-amino acid, each of the transport systems tested is stereospecific for the l-amino acid isomer. Apparent Michaelis constants for α-glycerol-P:dichloroindophenol reductase and α-glycerol-P oxidation are identical in intact and sonically disrupted membrane vesicles. Moreover, a similar apparent Michaelis constant is obtained for α-glycerol-P with respect to stimulation of amino acid transport. These findings indicate the α-glycerol-P oxidation per se is the rate-limiting step for amino acid transport in these preparations.