Increased synthesis of phosphatidylserine decarboxylase in a strain of Escherichia coli bearing a hybrid plasmid. Altered association of enzyme with the membrane.

A strain of Escherichia coli bearing a hybrid plasmid containing the psd gene, starved for isoleucine by the addition of valine, produces amounts of phosphatidyl-serine decarboxylase, a membrane-bound enzyme, about 40-fold higher than wild type. At least 98% of the enzyme from cells with high levels of decarboxylase is isolated in the inner, cytoplasmic membrane fraction if the cells are broken by osmotic lysis of spheroplasts following treatment with lysozyme/EDTA. In contrast, if cells containing these large amounts of enzyme are disrupted by sonication, 40 to 45% of the activity is recovered in the 100,000 times g supernatant fraction, whereas with wild type cells, only 5 to 10% is recovered in this fraction. About half of the decarboxylase in membranes saturated with the enzyme is thus only loosely bound, and readily removed by sonication, but not by osmotic lysis. This apparent saturation of the membrane with decarboxylase seems specific, since two other membrane-bound enzymes, phosphatidyl-glycerophosphate synthetase, and CDP-diglyceride synthetase, are not displaced into the supernatant fraction upon sonication. Fractionation on columns of agarose and by centrifugation through gradients of sucrose revealed that the decarboxylase in the supernatant is associated with lipid, in a complex with an apparent molecular weight of at least 5 times 10(6).