CpgD is a phosphoglycerate cytidylyltransferase required for ceramide diphosphoglycerate synthesis

Abstract

Lipopolysaccharide (LPS) is essential in most Gram-negative bacteria, but mutants of several species have been isolated that can survive in its absence. Caulobacter crescentus viability in the absence of LPS is partially dependent on the anionic sphingolipid ceramide diphosphoglycerate (CPG2). Genetic analyses showed that ccna_01210, which encodes a nucleotidyltransferase, is required for CPG2 production. Using purified recombinant protein, we determined that CCNA_01210 (CpgD) is a phosphoglycerate cytidylyltransferase which uses CTP and 3-phosphoglycerate to produce CDP-glycerate, which we hypothesize is the phosphoglycerate donor for CPG2 synthesis. CpgD had optimum activity at pH 7.5-8 in the presence of magnesium. CpgD exhibited Michaelis-Menten kinetics with respect to 3-phosphoglycerate (Km,app = 10.9 {+/-} 0.7 mM; Vmax,app = 0.72 {+/-} 0.02 {micro}mol/min/mg enzyme) and CTP (Km,app = 4.8 {+/-} 0.9 mM; Vmax,app = 0.44 {+/-} 0.03 {micro}mol/min/mg enzyme). The characterization of this enzyme uncovers another piece of the pathway towards CPG2 synthesis.