title: Structural Insight into Substrate Selection and Catalysis of Lipid Phosphate Phosphatase PgpB in the Cell Membrane.
abstract: PgpB belongs to the lipid phosphate phosphatase protein family and is one of three bacterial integral membrane phosphatases catalyzing dephosphorylation of phosphatidylglycerol phosphate (PGP) to generate phosphatidylglycerol. Although the structure of its apo form became recently available, the mechanisms of PgpB substrate binding and catalysis are still unclear. We found that PgpB was inhibited by phosphatidylethanolamine (PE) in a competitive mode in vitro Here we report the crystal structure of the lipid-bound form of PgpB. The structure shows that a PE molecule is stabilized in a membrane-embedded tunnel formed by TM3 and the "PSGH" fingerprint peptide near the catalytic site, providing structural insight into PgpB substrate binding mechanism. Noteworthy, in silico docking of varied lipid phosphates exhibited similar substrate binding modes to that of PE, and the residues in the lipid tunnel appear to be important for PgpB catalysis. The catalytic triad in the active site is essential for dephosphorylating substrates lysophosphatidic acid, phosphatidic acid, or sphingosine-1-phosphate but surprisingly not for the native substrate PGP. Remarkably, residue His-207 alone is sufficient to hydrolyze PGP, indicating a specific catalytic mechanism for PgpB in PG biosynthesis. We also identified two novel sensor residues, Lys-93 and Lys-97, on TM3. Our data show that Lys-97 is essential for the recognition of lyso-form substrates. Modification at the Lys-93 position may alter substrate specificity of lipid phosphate phosphatase proteins in prokaryotes versus eukaryotes. These studies reveal new mechanisms of lipid substrate selection and catalysis by PgpB and suggest that the enzyme rests in a PE-stabilized state in the bilayer.
authors.affiliation: From the Center for Membrane Biology, Department of Biochemistry and Molecular Biology, University of Texas Houston Medical School, Houston, Texas 77030 and.
authors.forename: Shuilong
authors.initials: S
authors.lastname: Tong
authors.affiliation: From the Center for Membrane Biology, Department of Biochemistry and Molecular Biology, University of Texas Houston Medical School, Houston, Texas 77030 and.
authors.forename: Yibin
authors.initials: Y
authors.lastname: Lin
authors.affiliation: From the Center for Membrane Biology, Department of Biochemistry and Molecular Biology, University of Texas Houston Medical School, Houston, Texas 77030 and.
authors.forename: Shuo
authors.initials: S
authors.lastname: Lu
authors.affiliation: Swiss Light Source, Paul Scherrer Institute, CH-5232 Villigen PSI, Switzerland.
authors.forename: Meitian
authors.initials: M
authors.lastname: Wang
authors.affiliation: From the Center for Membrane Biology, Department of Biochemistry and Molecular Biology, University of Texas Houston Medical School, Houston, Texas 77030 and.
authors.forename: Mikhail
authors.initials: M
authors.lastname: Bogdanov
authors.affiliation: From the Center for Membrane Biology, Department of Biochemistry and Molecular Biology, University of Texas Houston Medical School, Houston, Texas 77030 and lei.zheng@uth.tmc.edu.
authors.forename: Lei
authors.initials: L
authors.lastname: Zheng
mesh_terms: D002384:Catalysis; D020134:Catalytic Domain; D002462:Cell Membrane / Q000201:enzymology* / Q000235:genetics / Q000378:metabolism; D004926:Escherichia coli / Q000201:enzymology* / Q000235:genetics; D029968:Escherichia coli Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D008246:Lysophospholipids / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D010711:Phosphatidate Phosphatase / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D013110:Sphingosine / Q000031:analogs & derivatives* / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D013379:Substrate Specificity
chemical_list: D029968:Escherichia coli Proteins; D008246:Lysophospholipids; C000608554:PgpB protein, E coli; C060506:sphingosine 1-phosphate; D010711:Phosphatidate Phosphatase; D013110:Sphingosine; C032881:lysophosphatidic acid
keywords: X-ray crystallography; lipid phosphate dephosphorylation; phosphatase; phosphatidylglycerol; phospholipid metabolism; protein-lipid interaction; substrate selectivity