Brain microsomes catalyze the acylation of lysophosphatidylcholine (lysoPtdCho) in the presence and absence of added CoA derivatives. The catalytic activity is distributed widely in various subcellular fractions from rat or bovine cerebral cortex as measured by the conversion of 1-[14C]palmitoyl-sn-glycero-3-phosphocholine to [14C]PtdCho. Analysis of this latter compound revealed that the dipalmitoyl derivative is the predominant molecular species, which is formed in this reaction by transacylation between two [14C]lysoPtdCho molecules. This lysoPtdCho: lysoPtdCho transacylation reaction was enhanced several-fold by the addition of oleoyl-CoA, which also is an effective donor of acyl groups in the acyl-CoA: lysoPtdCho acyltransferase-catalyzed reaction. Measurements of the initial velocity of the transacylation reaction were used to determine kinetic constants. Apparent Km values for lysoPtdCho in the presence and absence of oleoyl-CoA were 29 microM and 104 microM, respectively, and the corresponding maximal velocities were 0.11 and 1.06 nmol.min-1.mg-1, respectively. Oleoyl-CoA at 4 microM produced half-maximal stimulation of the transacylation reaction. CoA also stimulated the rate of conversion of [14C]lysoPtdCho to [14C]PtdCho, either in the presence or absence of oleoyl-CoA, with a half-maximal effect of CoA at 80 microM. These results may be important in understanding the regulation of PtdCho synthesis and the mechanism by which acyl group composition of this compound is controlled.