poxB

Summary

Gene Symbol: poxB
Description: pyruvate dehydrogenase, thiamine triphosphate-binding, FAD-binding
Alias: ECK0862, JW0855
Species: Escherichia coli str. K-12 substr. MG1655
Products:     poxB

Top Publications

  1. Grabau C, Chang Y, Cronan J. Lipid binding by Escherichia coli pyruvate oxidase is disrupted by small alterations of the carboxyl-terminal region. J Biol Chem. 1989;264:12510-9 pubmed
    ..We report mutations generated by chemical and oligonucleotide-mediated site-directed mutagenesis of the poxB gene which result in enzymes defective in lipid activation...
  2. Chang Y, Cronan J. Sulfhydryl chemistry detects three conformations of the lipid binding region of Escherichia coli pyruvate oxidase. Biochemistry. 1997;36:11564-73 pubmed
    ..experiments detected a conformational change within the C-terminal segment of Escherichia coli pyruvate oxidase (PoxB), a lipid-activated homotetrameric enzyme, upon substrate binding [Chang, Y.-Y., & Cronan, J. E., Jr. (1995) J...
  3. O brien T, Blake R, Gennis R. Regulation by lipids of cofactor binding to a peripheral membrane enzyme: binding of thiamin pyrophosphate to pyruvate oxidase. Biochemistry. 1977;16:3105-9 pubmed
  4. Schrock H, Gennis R. High affinity lipid binding sites on the peripheral membrane enzyme pyruvate oxidase. Specific ligand effects on detergent binding. J Biol Chem. 1977;252:5990-5 pubmed
  5. Koland J, Gennis R. Identification of an active site cysteine residue in Escherichia coli pyruvate oxidase. J Biol Chem. 1982;257:6023-7 pubmed
    ..It is shown that methylmethanethiosulfonate and N-ethylmaleimide react with the same cysteine residue. Thus, the N-ethylmaleimide-sensitive residue is probably not directly involved in catalysis. ..
  6. Zhang T, Hager L. A single-step large-scale purification of pyruvate oxidase. Arch Biochem Biophys. 1987;257:485-7 pubmed
    ..It appears likely that this purification procedure can be used successfully with the family of enzymes which respond to both lipid and protease activation. ..
  7. Recny M, Hager L. Reconstitution of native Escherichia coli pyruvate oxidase from apoenzyme monomers and FAD. J Biol Chem. 1982;257:12878-86 pubmed
    ..The subsequent formation of native tetramers appears to proceed via an essentially irreversible dimer assembly pathway. ..
  8. Chang Y, Cronan J. An Escherichia coli mutant deficient in pyruvate oxidase activity due to altered phospholipid activation of the enzyme. Proc Natl Acad Sci U S A. 1984;81:4348-52 pubmed
    ..The mutant enzyme functions poorly in vivo, indicating that activation of the oxidase by phospholipids plays an important physiological role. ..
  9. Bertagnolli B, Hager L. Role of flavin in acetoin production by two bacterial pyruvate oxidases. Arch Biochem Biophys. 1993;300:364-71 pubmed
    ..The results indicate that the role for FAD in acetoin formation with both enzymes is based on a structural requirement and that FAD does not participate in a redox function in the acetoin reaction. ..

More Information

Publications70

  1. Chang Y, Wang A, Cronan J. Molecular cloning, DNA sequencing, and biochemical analyses of Escherichia coli glyoxylate carboligase. An enzyme of the acetohydroxy acid synthase-pyruvate oxidase family. J Biol Chem. 1993;268:3911-9 pubmed
    ..coli AHAS isozyme large and small subunits, Orf258 does not function as a Gcl subunit. Moreover, disruption of the chromosomal copy of orf258 did not affect growth on glyoxylate or glycolate. ..
  2. Blake R, O brien T, Gennis R, Hager L. Role of the divalent metal cation in the pyruvate oxidase reaction. J Biol Chem. 1982;257:9605-11 pubmed
    ..Both EPR and steady state kinetics data indicated complex formation with a dissociation constant of about 70 mM. ..
  3. Recny M, Grabau C, Cronan J, Hager L. Characterization of the alpha-peptide released upon protease activation of pyruvate oxidase. J Biol Chem. 1985;260:14287-91 pubmed
    ..From these analyses, we propose that the alpha-peptide may play a role in binding pyruvate oxidase to cell membrane phospholipids in vivo. ..
  4. Carter K, Gennis R. Reconstitution of the Ubiquinone-dependent pyruvate oxidase system of Escherichia coli with the cytochrome o terminal oxidase complex. J Biol Chem. 1985;260:10986-90 pubmed
    ..This is apparently not the case. It now is clear that both of the E. coli terminal oxidases act as ubiquinol-8 oxidases and, thus, ubiquinone-8 is the branch point between the two respiratory chains. ..
  5. Kiuchi K, Hager L. Reconstitution of the lipid-depleted pyruvate oxidase system of Escherichia coli: the palmitic acid effect. Arch Biochem Biophys. 1984;233:776-84 pubmed
    ..The stimulation of the electron transfer reaction by fatty acids occurs at the ubiquinone level of the electron transport chain. Ubiquinone-30 is rapidly reduced by pyruvate oxidase only in the presence of palmitic acid. ..
  6. Torchut E, Bourdillon C, Laval J. Reconstitution of a functional electron transfer in a biomimetic structure, including an electrode interface, phospholipid and ubiquinone molecules, and a membrane enzyme. Ann N Y Acad Sci. 1995;750:112-5 pubmed
  7. Chang Y, Cronan J, Li S, Reed K, Vanden Boom T, Wang A. Locations of the lip, poxB, and ilvBN genes on the physical map of Escherichia coli. J Bacteriol. 1991;173:5258-9 pubmed
  8. Abdel Hamid A, Attwood M, Guest J. Pyruvate oxidase contributes to the aerobic growth efficiency of Escherichia coli. Microbiology. 2001;147:1483-98 pubmed
    The metabolic importance of pyruvate oxidase (PoxB), which converts pyruvate directly to acetate and CO(2), was assessed using an isogenic set of genetically engineered strains of Escherichia coli...
  9. Schrock H, Gennis R. Specific ligand enhancement of the affinity of E. coli pyruvate oxidase for dipalmitoyl phosphatidylcholine. Biochim Biophys Acta. 1980;614:215-20 pubmed
    ..It is clearly shown that the protein-lipid interaction is dramatically modulated by the ligands bound at the catalytic active site on the enzyme and/or by the oxidation-reduction state of the flavin. ..
  10. Chang Y, Cronan J. Conversion of Escherichia coli pyruvate oxidase to an 'alpha-ketobutyrate oxidase'. Biochem J. 2000;352 Pt 3:717-24 pubmed
    Escherichia coli pyruvate oxidase (PoxB), a lipid-activated homotetrameric enzyme, is active on both pyruvate and 2-oxobutanoate ('alpha-ketobutyrate'), although pyruvate is the favoured substrate...
  11. Chang T, Ruan K. Exploration of pressure-induced dissociation of pyruvate oxidase. Cell Mol Biol (Noisy-le-grand). 2004;50:323-8 pubmed
  12. Weber A, Kögl S, Jung K. Time-dependent proteome alterations under osmotic stress during aerobic and anaerobic growth in Escherichia coli. J Bacteriol. 2006;188:7165-75 pubmed
    ..Eleven additional proteins (OtsB, YceI, YciE, YciF, YgaU, YjbJ, AcnA, MetL, PoxB, Ssb, and YhbO) were induced by osmotic stress imposed by NaCl...
  13. Chang Y, Cronan J. Genetic and biochemical analyses of Escherichia coli strains having a mutation in the structural gene (poxB) for pyruvate oxidase. J Bacteriol. 1983;154:756-62 pubmed
    ..The poxB locus was mapped by using closely linked transposon insertions to min 18.7 of the E...
  14. Recny M, Hager L. Isolation and characterization of the protease-activated form of pyruvate oxidase. Evidence for a conformational change in the environment of the flavin prosthetic group. J Biol Chem. 1983;258:5189-95 pubmed
    ..These results suggest that cleavage and release of the alpha peptide opens up the flavin active site and may be directly responsible for the observed stimulation of enzymatic activity. ..
  15. Cunningham C, Hager L. Crystalline pyruvate oxidase from Escherichia coli. II. Activation by phospholipids. J Biol Chem. 1971;246:1575-82 pubmed
  16. Marschall C, Hengge Aronis R. Regulatory characteristics and promoter analysis of csiE, a stationary phase-inducible gene under the control of sigma S and the cAMP-CRP complex in Escherichia coli. Mol Microbiol. 1995;18:175-84 pubmed
    ..The promoter region of poxB, the structural gene for pyruvate oxidase, which is also under the control of sigma S and cAMP-CRP, is very similar ..
  17. O brien T, Gennis R. Studies of the thiamin pyrophosphate binding site of Escherichia coli pyruvate oxidase. Evidence for an essential tryptophan residue. J Biol Chem. 1980;255:3302-7 pubmed
  18. O brien T, Shelton E, Mather M, Gennis R. Conformational studies of Escherichia coli pyruvate oxidase. Biochim Biophys Acta. 1982;705:321-9 pubmed
    ..The results indicate that the conformational changes in the protein due either to reduction of the flavin or to the binding of lipid activators are localized. ..
  19. Lin H, Bennett G, San K. Genetic reconstruction of the aerobic central metabolism in Escherichia coli for the absolute aerobic production of succinate. Biotechnol Bioeng. 2005;89:148-56 pubmed
    ..Mutations in the tricarboxylic acid cycle (sdhAB, icd, iclR) and acetate pathways (poxB, ackA-pta) of E. coli were created to construct the glyoxylate cycle for aerobic succinate production...
  20. Tittmann K. Reaction mechanisms of thiamin diphosphate enzymes: redox reactions. FEBS J. 2009;276:2454-68 pubmed publisher
    ..Viable mechanisms of reductive acetylation in pyruvate dehydrogenase multi-enzyme complex, and of electron transfer in the peripheral membrane enzyme pyruvate oxidase from Escherichia coli, are also discussed. ..
  21. Wang A, Chang Y, Cronan J. Role of the tetrameric structure of Escherichia coli pyruvate oxidase in enzyme activation and lipid binding. J Biol Chem. 1991;266:10959-66 pubmed
    ..We found that when two poxB gene alleles coexisted in cells, heterotetrameric species were formed in addition to homotetramers...
  22. Jantama K, Zhang X, Moore J, Shanmugam K, Svoronos S, Ingram L. Eliminating side products and increasing succinate yields in engineered strains of Escherichia coli C. Biotechnol Bioeng. 2008;101:881-93 pubmed publisher
    ..that disrupt fermentation pathways for alternative products (ldhA::FRT, adhE::FRT, ackA::FRT, focA-pflB::FRT, mgsA, poxB) with growth-based selection for increased ATP production. The resulting strain, KJ073, produced 1...
  23. Houghton R, Swoboda B. Kinetics of the mechanism of action of flavin pyruvate oxidase from an acetate requiring mutant of Escherichia coli. FEBS Lett. 1973;30:277-80 pubmed
  24. Chang Y, Wang A, Cronan J. Expression of Escherichia coli pyruvate oxidase (PoxB) depends on the sigma factor encoded by the rpoS(katF) gene. Mol Microbiol. 1994;11:1019-28 pubmed
    The activity of Escherichia coli pyruvate oxidase (PoxB) was shown to be growth-phase dependent; the enzyme activity reaches a maximum at early stationary phase...
  25. Schrock H, Gennis R. The binding of a fluorescent activator 2-(N-decyl)aminonaphthalene-6-sulfonic acid to pyruvate oxidase. Biochim Biophys Acta. 1980;615:10-8 pubmed
    ..However, in both these cases, the amount of bound detergent was nearly an order of magnitude less than the values obtained using DNS. ..
  26. Chang Y, Cronan J. Molecular cloning, DNA sequencing, and enzymatic analyses of two Escherichia coli pyruvate oxidase mutants defective in activation by lipids. J Bacteriol. 1986;167:312-8 pubmed
    ..The results of the amino acid substitutions in the mutant proteins, leading to the functional alteration of the enzyme, are discussed. ..
  27. Stevens D, Gennis R. Studies on the quaternary structure of Escherichia coli pyruvate oxidase. J Biol Chem. 1980;255:379-83 pubmed
    ..A photoreactive, nonspecific probe, 1-azidonaphthalene 5-sulfonate, was used to label the protein surface with an extrinsic fluorophore. It is concluded that under steady state assay conditions the oxidase remains tetrameric. ..
  28. Mather M, Schopfer L, Massey V, Gennis R. Studies of the flavin adenine dinucleotide binding region in Escherichia coli pyruvate oxidase. J Biol Chem. 1982;257:12887-92 pubmed
    ..Restrictions to the accessibility of the active site of the enzyme are suggested by the fact that sulfite does not form an adduct with the flavin in the native enzyme. ..
  29. Moreau P. Diversion of the metabolic flux from pyruvate dehydrogenase to pyruvate oxidase decreases oxidative stress during glucose metabolism in nongrowing Escherichia coli cells incubated under aerobic, phosphate starvation conditions. J Bacteriol. 2004;186:7364-8 pubmed
    ..Inactivation of pyruvate oxidase (PoxB) activity decreased the production of acetate by half and significantly increased the production of TBARS...
  30. Torchut E, Bourdillon C, Laval J. Reconstitution of functional electron transfer between membrane biological elements in a two-dimensional lipidic structure at the electrode interface. Biosens Bioelectron. 1994;9:719-23 pubmed
    ..A membrane enzyme, pyruvate oxidase from E. coli, was successfully incorporated into this artificial bilayer and was found to be able to exchange electrons with ubiquinone present in the bilayer. ..
  31. Flores N, de Anda R, Flores S, Escalante A, Hernandez G, Martinez A, et al. Role of pyruvate oxidase in Escherichia coli strains lacking the phosphoenolpyruvate:carbohydrate phosphotransferase system. J Mol Microbiol Biotechnol. 2004;8:209-21 pubmed
    ..Inactivation of poxB caused a severe reduction in growth rate in strain PB11 when grown in the fermentor with medium containing glucose ..
  32. Russell P, Schrock H, Gennis R. Lipid activation and protease activation of pyruvate oxidase. Evidence suggesting a common site of interaction on the protein. J Biol Chem. 1977;252:7883-7 pubmed
  33. Koland J, Miller M, Gennis R. Reconstitution of the membrane-bound, ubiquinone-dependent pyruvate oxidase respiratory chain of Escherichia coli with the cytochrome d terminal oxidase. Biochemistry. 1984;23:445-53 pubmed
    ..coli aerobic respiratory chain. These investigations suggest a relatively simple organization for the E. coli respiratory chain. ..
  34. Mather M, Gennis R. Kinetic studies of the lipid-activated pyruvate oxidase flavoprotein of Escherichia coli. J Biol Chem. 1985;260:16148-55 pubmed
    ..Activation appears to result in both increased rates of electron transfer into and out of the flavin. ..
  35. Weidner A, Neumann P, Wille G, Stubbs M, Tittmann K. Crystallization and preliminary X-ray diffraction analysis of full-length and proteolytically activated pyruvate oxidase from Escherichia coli. Acta Crystallogr Sect F Struct Biol Cryst Commun. 2008;64:179-81 pubmed publisher
  36. Neumann P, Weidner A, Pech A, Stubbs M, Tittmann K. Structural basis for membrane binding and catalytic activation of the peripheral membrane enzyme pyruvate oxidase from Escherichia coli. Proc Natl Acad Sci U S A. 2008;105:17390-5 pubmed publisher
    ..The isolated C terminus, which has no intrinsic helix propensity, folds into a helical structure in the presence of micelles. ..
  37. Raj T, Russell P, Flygare W, Gennis R. Quasi-elastic light scattering studies on pyruvate oxidase. Biochim Biophys Acta. 1977;481:42-9 pubmed
    ..The sedimentation velocity-diffusion technique was used to obtain the molecular weight of this tetrameric enzyme, about 252 000 with a value of f/f0 of 1.25. ..
  38. O brien T, Gennis R. Studies on the interaction between Escherichia coli pyruvate oxidase and a detergent activator by utilization of the fluorescence probe bis(8-p-toluidino-1-naphthalenesulfonate). Biochemistry. 1979;18:804-9 pubmed
  39. Bertagnolli B, Hager L. Activation of Escherichia coli pyruvate oxidase enhances the oxidation of hydroxyethylthiamin pyrophosphate. J Biol Chem. 1991;266:10168-73 pubmed
    ..Alternatively, the rate-limiting step in the overall reaction with the activated enzyme shifts to one of the partial steps in the decarboxylation reaction. ..
  40. Williams F, Hager L. Crystalline flavin pyruvate oxidase from Escherichia coli. I. Isolation and properties of the flavoprotein. Arch Biochem Biophys. 1966;116:168-76 pubmed
  41. Mather M, Gennis R. Spectroscopic studies of pyruvate oxidase flavoprotein from Escherichia coli trapped in the lipid-activated form by cross-linking. J Biol Chem. 1985;260:10395-7 pubmed
  42. Barassi C, Kranz R, Gennis R. Characterization of monoclonal antibodies directed against pyruvate oxidase from Escherichia coli: modulation of antibody-induced inhibition by enzyme conformation. Biochem Biophys Res Commun. 1986;137:884-91 pubmed
    ..Hence, the coupling between antibody binding and the conformation at the active site can itself be modulated by the conformation of the protein. ..
  43. Cunningham C, Hager L. Crystalline pyruvate oxidase from Escherichia coli. 3. Phospholipid as an allosteric effector for the enzyme. J Biol Chem. 1971;246:1583-9 pubmed
  44. Chang Y, Cronan J. Detection by site-specific disulfide cross-linking of a conformational change in binding of Escherichia coli pyruvate oxidase to lipid bilayers. J Biol Chem. 1995;270:7896-901 pubmed
    ..The cross-linked oxidase is enzymatically active and remains able to associate with lipid micelles. These results argue strongly that lipid bilayer binding of pyruvate oxidase involves pairing of the C termini of two subunits. ..
  45. Moreau P. The lysine decarboxylase CadA protects Escherichia coli starved of phosphate against fermentation acids. J Bacteriol. 2007;189:2249-61 pubmed
    ..P(i)-starved cells resulted primarily from the catabolism of glucose into organic acids through pyruvate oxidase (PoxB) and pyruvate-formate lyase (PflB) under aerobic and anaerobic conditions, respectively...
  46. Grabau C, Cronan J. In vivo function of Escherichia coli pyruvate oxidase specifically requires a functional lipid binding site. Biochemistry. 1986;25:3748-51 pubmed
    ..Strains producing the truncated protein were devoid of oxidase activity in vivo. This result indicates that binding to membrane lipids is specifically required for function of the oxidase in vivo; activation alone does not suffice. ..
  47. Marchal D, Pantigny J, Laval J, Moiroux J, Bourdillon C. Rate constants in two dimensions of electron transfer between pyruvate oxidase, a membrane enzyme, and ubiquinone (coenzyme Q8), its water-insoluble electron carrier. Biochemistry. 2001;40:1248-56 pubmed
    ..The kinetic constants we found show that the activated enzyme can be efficiently regulated by the oxidation level of the quinone pool in natural membranes. ..
  48. Cunningham C, Hager L. Reactivation of the lipid-depleted pyruvate oxidase system from Escherichia coli with cell envelope neutral lipids. J Biol Chem. 1975;250:7139-46 pubmed
    ..In fact, these compounds appear to suppress restoration of activity when they are added to reaction mixtures containing neutral lipid and phospholipase-treated particles. ..
  49. O brien T, Kluger R, Pike D, Gennis R. Phosphonate analogues of pyruvate. Probes of substrate binding to pyruvate oxidase and other thiamin pyrophosphate-dependent decarboxylases. Biochim Biophys Acta. 1980;613:10-7 pubmed
    ..Kinetic data suggest that lipid activators of pyruvate oxidase increase the affinity of the enzyme for pyruvate and its analogues. ..
  50. Houghton R. The effect of temperature, urea and N-ethylmaleimide on pyruvate oxidase (EC 1.2.2.2) activity. Int J Biochem. 1979;10:205-8 pubmed
  51. Russell P, Hager L, Gennis R. Characterization of the proteolytic activation of pyruvate oxidase. Control by specific ligands and by the flavin oxidation-reduction state. J Biol Chem. 1977;252:7877-82 pubmed
  52. Grabau C, Cronan J. Molecular cloning of the gene (poxB) encoding the pyruvate oxidase of Escherichia coli, a lipid-activated enzyme. J Bacteriol. 1984;160:1088-92 pubmed
    The pyruvate oxidase structural gene (poxB) of Escherichia coli was cloned into derivatives of plasmid pBR322...
  53. Grabau C, Cronan J. Nucleotide sequence and deduced amino acid sequence of Escherichia coli pyruvate oxidase, a lipid-activated flavoprotein. Nucleic Acids Res. 1986;14:5449-60 pubmed
    The entire nucleotide sequence of the poxB (pyruvate oxidase) gene of Escherichia coli K-12 has been determined by the dideoxynucleotide (Sanger) sequencing of fragments of the gene cloned into a phage M13 vector...
  54. Koland J, O brien T, Gennis R. Role of arginine in the binding of thiamin pyrophosphate to Escherichia coli pyruvate oxidase. Biochemistry. 1982;21:2656-600 pubmed
    ..Furthermore, inactivation by 2,3-butanedione is attended by at least a 100-fold decrease in affinity of the enzyme for TPP. These results suggest a direct role for arginine in the binding of the cofactor. ..
  55. O brien T, Schrock H, Russell P, Blake R, Gennis R. Preparation of Escherichia coli pyruvate oxidase utilizing a thiamine pyrophosphate affinity column. Biochim Biophys Acta. 1976;452:13-29 pubmed
    ..The enzyme is a tetramer with a subunit molecular weight of approx. 60 000. The subunits appear to be identical. The isoelectric point of pyruvate oxidase is 5.6. ..
  56. Bertagnolli B, Hager L. Minimum requirements for protease activation of flavin pyruvate oxidase. Biochemistry. 1991;30:8131-7 pubmed
    ..Endoproteinase Glu-C specificity predicts hydrolytic cleavage of the peptide bond between Glu-561 and Val-562 with the removal of 11 residues from the carboxy terminus of the enzyme. ..
  57. Vemuri G, Minning T, Altman E, Eiteman M. Physiological response of central metabolism in Escherichia coli to deletion of pyruvate oxidase and introduction of heterologous pyruvate carboxylase. Biotechnol Bioeng. 2005;90:64-76 pubmed
    ..The presence of PYC in a poxB mutant resulted in a 42% lower maintenance coefficient and a 42% greater biomass yield compared to the wildtype...
  58. Van Dyk T, Ayers B, Morgan R, LaRossa R. Constricted flux through the branched-chain amino acid biosynthetic enzyme acetolactate synthase triggers elevated expression of genes regulated by rpoS and internal acidification. J Bacteriol. 1998;180:785-92 pubmed
    ..open reading frames, f253a, f415, frvX, o513, o521, yciG, and yohF, and five known genes, inaA, IdcC, osmY, poxB, and sohA...
  59. Koland J, Gennis R. Proximity of reactive cysteine residue and flavin in Escherichia coli pyruvate oxidase as estimated by fluorescence energy transfer. Biochemistry. 1982;21:4438-42 pubmed
    ..Thus, the TPP binding site and the FAD coenzyme are likely in close proximity. ..
  60. Hamilton S, Recny M, Hager L. Identification of the high-affinity lipid binding site in Escherichia coli pyruvate oxidase. Biochemistry. 1986;25:8178-83 pubmed
    ..The present work locates the lipid binding site very close to this peptide. The significance of these results for the mechanism of activation of pyruvate oxidase and other lipid-activated systems is discussed. ..
  61. Chang Y, Cronan J. Common ancestry of Escherichia coli pyruvate oxidase and the acetohydroxy acid synthases of the branched-chain amino acid biosynthetic pathway. J Bacteriol. 1988;170:3937-45 pubmed