Effect of phospholipids on the activity of sialosyl lactosylceramide (GM3): N-acetylgalactosaminyl transferase from chick embryo brain

Abstract

A preparation of UDP-GalNAc:sialosyl lactosylceramide N-acetylgalactosaminyl transferase (E.C. 2.4.1.92) obtained from 14-day chick embryo brain was delipidated partially by treatment with cold acetone in the presence of varying amounts of sodium dodecyl sulphate (SDS). The lipid content and the enzyme activity of the preparation decreased as the concentration of SDS increased. At 0.3% SDS the lipid content was about 30% and the enzyme activity about 15% of the original. The activity could be restored up to 60% of the original by added phospholipids, provided the removal of endogenous lipids did not exceed 70%. Phospholipids with different composition showed different abilities to restore the enzyme activity. Among phosphatidylcholines the decreasing order of effectivity was dilauroyl----dimiristoyl----dipalmitoyl----distearoyl-choline. Dimiristoyl phosphatidylcholine, dimiristoyl phosphatidylglycerol and dipalmitoyl phosphatidylglycerol activated the enzyme more effectively than dimiristoyl phosphatidyl ethanolamine, dimiristoyl phosphatidic acid or brain phosphatidylserine. No correlation was found between the activating ability and the charge in the polar head group of the lipid added. Addition of dilauroyl phosphatidylcholine to the delipidated preparation increased about 5-fold the Vmax without affecting the apparent Km for both the donor nucleotide and acceptor glycolipid. The data suggest that the lipid composition of the enzyme environment constitutes a potential level of regulation of the activity of this key enzyme of ganglioside biosynthesis.