Topology and active site of PlsY - The bacterial acylphosphate: Glycerol-3-phosphate acyltransferase

被引:31
作者
Lu, Ying-Jie
Zhang, Fan
Grimes, Kimberly D.
Lee, Richard E.
Rock, Charles O.
机构
[1] St Jude Childrens Res Hosp, Dept Infect Dis, Memphis, TN 38105 USA
[2] Univ Tennessee, Ctr Hlth Sci, Dept Pharmaceut Sci, Memphis, TN 38163 USA
关键词
D O I
10.1074/jbc.M700374200
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
The most widely distributed biosynthetic pathway to initiate phosphatidic acid formation in bacterial membrane phospholipid biosynthesis involves the conversion of acyl-acyl carrier protein to acylphosphate by PlsX and the transfer of the acyl group from acylphosphate to glycerol 3-phosphate by an integral membrane protein, PlsY. The membrane topology of Streptococcus pneumoniae PlsY was determined using the substituted cysteine accessibility method. PlsY has five membrane-spanning segments with the amino terminus and two short loops located on the external face of the membrane. Each of the three larger cytoplasmic domains contains a highly conserved sequence motif. Site-directed mutagenesis revealed that each conserved domain was critical for PlsY catalysis. Motif 1 had an essential serine and arginine residue. Motif 2 had the characteristics of a phosphate-binding loop. Mutations of the conserved glycines in motif 2 to alanines resulted in a K-m defect for glycerol 3-phosphate binding leading to the conclusion that this motif corresponded to the glycerol 3-phosphate binding site. Motif 3 contained a conserved histidine and asparagine that were important for activity and a glutamate that was critical to the structural integrity of PlsY. PlsY was noncompetitively inhibited by palmitoyl-CoA. These data define the membrane architecture and the critical active site residues in the PlsY family of bacterial acyltransferases.
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页码:11339 / 11346
页数:8
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