Effects of bilayer thickness on the activity of diacylglycerol kinase of Escherichia coli

We have developed a procedure for the reconstitution of Escherichia coli di acylglycerol kinase (DGK) into phospholipid bilayers containing diacylglyce rol substrate. When DC;K is reconstituted into a series of phosphatidylchol ines containing monounsaturated fatty acyl chains, activity against dihexan oyl-glycerol (DHG) as a substrate was found to be markedly dependent on the fatty acyl chain length with the highest activity in dioleoylphosphatidylc holine [di(C18:1)PC] and a lower activity in bilayers with shorter or longe r fatty acyl chains. Low activities in the short chain phospholipid dimyris toleoylphosphatidylcholine [di(C14:1)PC] followed from an increase in the K -m value for DHG and ATP, with no effect on v(max). In contrast, in the lon g chain lipid dierucoylphosphatidylcholine [di(C34:1)PC], the low activity followed from a decrease in v(max) with no effect on K-m. In mixtures of tw o phosphatidylcholines with different chain lengths, the activity correspon ded to that expected for the average chain length of the mixture. Cholester ol increased the activity in di(C14:1)PC but slightly decreased it in di(C1 8:1)PC or di(C24:1)PC, effects that could follow from changes in bilayer th ickness caused by cholesterol.