Publications

• The phosphonopyruvate decarboxylase from Bacteroides fragilis.

  Zhang G., Dai J., Lu Z., Dunaway-Mariano D.

  The Bacteroides fragilis capsular polysaccharide complex is the major virulence factor for abscess formation in human hosts. Polysaccharide B of this complex contains a 2-aminoethylphosphonate functional group. This functional group is synthesized in three steps, one of which is catalyzed by phosp ... >> More

  The Bacteroides fragilis capsular polysaccharide complex is the major virulence factor for abscess formation in human hosts. Polysaccharide B of this complex contains a 2-aminoethylphosphonate functional group. This functional group is synthesized in three steps, one of which is catalyzed by phosphonopyruvate decarboxylase. In this paper, we report the cloning and overexpression of the B. fragilis phosphonopyruvate decarboxylase gene (aepY), purification of the phosphonopyruvate decarboxylase recombinant protein, and the extensive characterization of the reaction that it catalyzes. The homotrimeric (41,184-Da subunit) phosphonopyruvate decarboxylase catalyzes (kcat = 10.2 +/-0.3 s-1) the decarboxylation of phosphonopyruvate (Km = 3.2 +/-0.2 microm) to phosphonoacetaldehyde (Ki = 15 +/-2 microm) and carbon dioxide at an optimal pH range of 7.0-7.5. Thiamine pyrophosphate (Km = 13 +/-2 microm) and certain divalent metal ions (Mg(II) Km = 82 +/-8 microm; Mn(II) Km = 13 +/- 1 microm; Ca(II) Km = 78 +/-6 microm) serve as cofactors. Phosphonopyruvate decarboxylase is a member of the alpha-ketodecarboxylase family that includes sulfopyruvate decarboxylase, acetohydroxy acid synthase/acetolactate synthase, benzoylformate decarboxylase, glyoxylate carboligase, indole pyruvate decarboxylase, pyruvate decarboxylase, the acetyl phosphate-producing pyruvate oxidase, and the acetate-producing pyruvate oxidase. The Mg(II) binding residue Asp-260, which is located within the thiamine pyrophosphate binding motif of the alpha-ketodecarboxylase family, was shown by site-directed mutagenesis to play an important role in catalysis. Pyruvate (kcat = 0.05 s-1, Km = 25 mm) and sulfopyruvate (kcat approximately 0.05 s-1; Ki = 200 +/-20 microm) are slow substrates for the phosphonopyruvate decarboxylase, indicating that this enzyme is promiscuous. << Less

  J Biol Chem 278:41302-41308(2003) [PubMed] [EuropePMC]

• Interaction of inhibitors with phosphoenolpyruvate mutase: implications for the reaction mechanism and the nature of the active site.

  Seidel H.M., Knowles J.R.

  The active site and mechanism of action of the enzyme phosphoenolpyruvate mutase have been probed using substrate and intermediate analogues as inhibitors of the mutase-catalyzed reaction. Smaller anions (e.g. sulfite, nitrate, phosphinate, and bicarbonate) are noncompetitive inhibitors of the mut ... >> More

  The active site and mechanism of action of the enzyme phosphoenolpyruvate mutase have been probed using substrate and intermediate analogues as inhibitors of the mutase-catalyzed reaction. Smaller anions (e.g. sulfite, nitrate, phosphinate, and bicarbonate) are noncompetitive inhibitors of the mutase, while larger anions in the complementary series (sulfate, phosphonate, phosphate) inhibit competitively. Combining oxalate, an intermediate analogue that is a potent inhibitor of the mutase (Ki = 25 microM), with small, noncompetitive inhibitor anions results in synergistic inhibition of the mutase, suggesting that the combined presence of oxalate and anion creates a "bimolecular transition-state analogue". The phosphoenolpyruvate (PEP) mutase genes from Tetrahymena and Streptomyces are known, and these enzymes share significant amino acid sequence similarity to the isocitrate lyase gene from Ricinus. Despite their seeming structural unrelatedness to the substrates of PEP mutase, several isocitrate analogues are good inhibitors, suggesting that isocitrate lyase and PEP mutase are evolutionarily related. An active-site model has been developed that is in accord with the data presented, which are consistent with a mechanism involving the intermediacy of a phosphoenzyme. << Less

  Biochemistry 33:5641-5646(1994) [PubMed] [EuropePMC]

  This publication is cited by 1 other entry.

• Studies on the biosynthesis of bialaphos. Biochemical mechanism of C-P bond formation: discovery of phosphonopyruvate decarboxylase which catalyzes the formation of phosphonoacetaldehyde from phosphonopyruvate.

  Nakashita H., Watanabe K., Hara O., Hidaka T., Seto H.

  The biosynthetic step following the phosphoenolpyruvate (PEP) phosphomutase reaction which forms a C-P bond of bialaphos was proven by the identification of phosphonopyruvate (PnPy) and phosphonoacetaldehyde (PnAA) as intermediates in the culture broth of Streptomyces hygroscopicus, a producing or ... >> More

  The biosynthetic step following the phosphoenolpyruvate (PEP) phosphomutase reaction which forms a C-P bond of bialaphos was proven by the identification of phosphonopyruvate (PnPy) and phosphonoacetaldehyde (PnAA) as intermediates in the culture broth of Streptomyces hygroscopicus, a producing organism of bialaphos, and by detection of enzymatic decarboxylation of PnPy to PnAA. Purified PnPy decarboxylase turned out to require thiamine diphosphate and Mg2+ as cofactors. PnPy decarboxylase drives the unfavorable forward reaction to form PnPy catalyzed by PEP phosphomutase and is suggested to be essential to C-P compound biosynthesis. << Less

  J. Antibiot. 50:212-219(1997) [PubMed] [EuropePMC]