Inactivation of the gene for phospholipid N-methyltransferase in Sinorhizobium meliloti: phosphatidylcholine is required for normal growth

In phosphatidylcholine (PC)-containing prokaryotes, only the methylation pa thway of PC biosynthesis was thought to occur. However, a second choline-de pendent pathway for PC formation, the PC synthase (Pcs) pathway, exists in Sinorhizobium (Rhizobium) meliloti in which choline is condensed with CDP-d iacylglyceride. Here, we characterize the methylation pathway of PC biosynt hesis in S. meliloti. A mutant deficient in phospholipid N-methyltransferas e (Pmt) was complemented with a S. meliloti gene bank and the complementing DNA was sequenced. A gene coding for a S-adenosylmethionine-dependent N-me thyltransferase was identified as the sinorhizobial Pmt, which showed littl e similarity to the corresponding enzyme from Rhodobacter sphaeroides. Upon expression of the sinorhizobial Pmt, besides phosphatidylcholine, the meth ylated intermediates of the methylation pathway, monomethylphosphatidyletha nolamine and dimethylphosphatidylethanolamine, are also formed. When Pmt-de ficient mutants of S. meliloti are grown on minimal medium, they cannot for m PC, and they grow significantly more slowly than the wild type. Growth of the Pmt-deficient mutant in the presence of choline allows for PC formatio n via the Pcs pathway and restores wild-type-like growth. Double knock-out mutants, deficient in Pmt and in Pcs, are unable to form PC and show reduce d growth even in the presence of choline. These results suggest that PC is required for normal growth of S. meliloti.