Lysophosphatidylcholine acyltransferase activity in Saccharomyces cerevisiae: regulation by a high-affinity Zn2+ binding site.

Saccharomyces cerevisiae cells were demonstrated to contain lysophosphatidylcholine (lysoPtdCho) acyltransferase (E.C. 2.3.1.23) activity. The enzyme displayed Km(app) of 69 microM for lysoPtdCho and 152 microM for oleoyl CoA. Enzyme activity was not affected by the addition of 1 mM Mg2+, Mn2+, Ca2+, or 200 mM EDTA. However, Zn2+ inhibited lysoPtdCho acyltransferase activity to 33% control values at 0.1 mM and to 7% at 1.0 mM Zn2+. To further explore the possibility that lysoPtdCho acyltransferase may contain a high-affinity Zn2+ binding site, we tested the strong Zn2+ chelator o-phenanthroline for its ability to inhibit enzyme activity. LysoPtdCho acyltransferase activity was inhibited to 18 and 27%, respectively, those of control values in the presence of 2 and 1 mM o-phenanthroline, implying that a high-affinity Zn2+ binding site exists in lysoPtdCho acyltransferase or in an accessory protein that is essential for protein stability and/or activity. Saccharomyces cerevisiae lysoPtdCho acyltransferase activity displayed a broad lysoPtdCho fatty acyl chain substrate specificity utilizing lysoPtdCho molecules ranging in length from C10-C20 (the entire range tested). In addition, the enzyme was capable of using the ether-linked analog of lysoPtdCho, 1-O-alkyl-2-hydroxy-sn-3-glycerophosphocholine, as a substrate. The ability of S. cerevisiae to incorporate radiolabeled 1-O-alkyl-2-hydroxy-sn-3-glycerophosphocholine into phosphatidylcholine in vitro was exploited to demonstrate a direct precursor-product relationship between lysoPtdCho molecules and their incorporation into phosphatidylcholine in vivo. Identical labeling results were obtained in S. cerevisiae cells disrupted for their major transacylase activity, PLB1, demonstrating that the incorporation of lysolipid was via acyltransferase, and not transacylase, activity.