sn-Glycerol-3-phosphate auxotrophy of plsB strains of Escherichia coli: evidence that a second mutation, plsX, is required

Abstract

sn-Glycerol-3-phosphate auxotrophs defective in phospholipid synthesis contain a Km-defective sn-glycerol-3-phosphate acyltransferase. Detailed genetic analysis revealed that two mutations were required for the auxotrophic phenotype. One mutation, in the previously described plsB locus (sn-glycerol-3-phosphate acyltransferase structural gene), mapped near min 92 on the Escherichia coli linkage map. Isolation of Tn10 insertions cotransducible with the auxotrophy in phage P1 crosses revealed that a second mutation was required with plsB26 to confer the sn-glycerol-3-phosphate auxotrophic phenotype. This second locus, plsX, mapped between pyrC and purB near min 24 on the E. coli linkage map. Tn10 insertions near plsX allowed detailed mapping of the genetic loci in this region. A clockwise gene order putA pyrC flbA flaL flaT plsX fabD ptsG thiK purB was inferred from results of two- and three-factor crosses. Strains harboring the four possible configurations of the mutant and wild-type plsB and plsX loci were constructed. Isogenic plsB+ plsX+, plsB+ plsX50, and plsB26 plsX+ strains grew equally well on glucose minimal medium without sn-glycerol-3-phosphate. In addition, plsX or plsX+ had no apparent effect on sn-glycerol-3-phosphate acyltransferase activity measured in membrane preparations. The molecular basis for the plsX requirement for conferral of sn-glycerol-3-phosphate auxotrophy in these strains remains to be established.