Cyclopropane fatty acyl synthases (CFA synthases) are
enzymes that catalyse the addition of a methylene group across cis double bonds of monounsaturated fatty acyl chains in
lipids. We have investigated the function of two putative genes, cfa1 and cfa2, proposed to code for CFA synthases in Sinorhizobium meliloti. Total
fatty acid composition and
fatty acid distributions within
lipid classes for wild-type and cfa1 and cfa2 mutant strains grown under Pi
starvation and in acidic culture conditions were obtained by GC/MS and by infusion ESI/MS/MS, respectively. For wild-type cells and the cfa1 mutant, total
cyclopropane fatty acids (CFAs) increased by 10% and 15% under Pi
starvation and acidic conditions, respectively; whereas in the cfa2 mutant, CFAs were less than 0.1% of wild-type under both growth conditions. Reporter gene fusion experiments revealed that cfa1 and cfa2 were expressed at similar levels in free-living cells. Thus under the conditions we examined, cfa2 was required for the cyclopropanation of
lipids in S. meliloti whereas the role of cfa1 remains to be determined. Analysis of intact
lipids revealed that cyclopropanation occurred on cis-11-octadecenoic
acid located in either the sn-1 or the sn-2 position in
phospholipids and that cyclopropanation in the sn-2 position occurred to a greater extent in
phosphatidylcholines and sulfoquinovosyldiacylglycerols under acidic conditions than under Pi
starvation. The cfa2 gene was also required for cyclopropanation of non-
phosphorus-containing
lipids. Principal components analysis revealed no differences in the cyclopropanation of four
lipid classes. We concluded that cyclopropanation occurred independently of the polar head group. Neither cfa1 nor cfa2 was required for symbiotic nitrogen fixation.