Publications

• Molecular characterization of Escherichia coli malate synthase G. Differentiation with the malate synthase A isoenzyme.

  Molina I., Badia J., Aguilar J.T., Pellicer M.T., Baldoma L.

  Two genes encoding the enzymes malate synthase G and glycolate oxidase, have been linked to locus glc (64.5 min), responsible for glycolate utilization in Escherichia coli. The gene encoding malate synthase G, for which we propose the notation glcB, has been cloned, sequenced and found to correspo ... >> More

  Two genes encoding the enzymes malate synthase G and glycolate oxidase, have been linked to locus glc (64.5 min), responsible for glycolate utilization in Escherichia coli. The gene encoding malate synthase G, for which we propose the notation glcB, has been cloned, sequenced and found to correspond to a 2262-nucleotide open-reading frame, which can encode a 723-amino-acid polypeptide, clearly different from the isoenzyme malate synthase A, which has 533 amino acids. Northern-blot experiments indicate that glcB was expressed as an apparently monocistronic transcript, inducible by glycolate. Malate synthase G was purified to near homogeneity. The molecular mass determined by gel filtration yielded a value of 82 kDa for the purified enzyme and the same value as for the crude extract enzyme, indicating a monomeric structure. Despite the lower sequence similarity between malate synthase G and the other reported malate synthases, three out of nine consensus boxes defined in most of these enzymes are conserved in addition to a cysteine residue that has been reported to be important for the catalytic mechanisms. << Less

  Eur. J. Biochem. 224:541-548(1994) [PubMed] [EuropePMC]

• Structure of the Escherichia coli malate synthase G:pyruvate:acetyl-coenzyme A abortive ternary complex at 1.95 A resolution.

  Anstrom D.M., Kallio K., Remington S.J.

  Malate synthase, an enzyme of the glyoxylate pathway, catalyzes the condensation and subsequent hydrolysis of acetyl-coenzyme A (acetyl-CoA) and glyoxylate to form malate and CoA. In the present study, we present the 1.95 A-resolution crystal structure of Escherichia coli malate synthase isoform G ... >> More

  Malate synthase, an enzyme of the glyoxylate pathway, catalyzes the condensation and subsequent hydrolysis of acetyl-coenzyme A (acetyl-CoA) and glyoxylate to form malate and CoA. In the present study, we present the 1.95 A-resolution crystal structure of Escherichia coli malate synthase isoform G in complex with magnesium, pyruvate, and acetyl-CoA, and we compare it with previously determined structures of substrate and product complexes. The results reveal how the enzyme recognizes and activates the substrate acetyl-CoA, as well as conformational changes associated with substrate binding, which may be important for catalysis. On the basis of these results and mutagenesis of active site residues, Asp 631 and Arg 338 are proposed to act in concert to form the enolate anion of acetyl-CoA in the rate-limiting step. The highly conserved Cys 617, which is immediately adjacent to the presumed catalytic base Asp 631, appears to be oxidized to cysteine-sulfenic acid. This can explain earlier observations of the susceptibility of the enzyme to inactivation and aggregation upon X-ray irradiation and indicates that cysteine oxidation may play a role in redox regulation of malate synthase activity in vivo. There is mounting evidence that enzymes of the glyoxylate pathway are virulence factors in several pathogenic organisms, notably Mycobacterium tuberculosis and Candida albicans. The results described in this study add insight into the mechanism of catalysis and may be useful for the design of inhibitory compounds as possible antimicrobial agents. << Less

  Protein Sci. 12:1822-1832(2003) [PubMed] [EuropePMC]

• Biochemical and structural studies of malate synthase from Mycobacterium tuberculosis.

  Smith C.V., Huang C.C., Miczak A., Russell D.G., Sacchettini J.C., Honer zu Bentrup K.

  Establishment or maintenance of a persistent infection by Mycobacterium tuberculosis requires the glyoxylate pathway. This is a bypass of the tricarboxylic acid cycle in which isocitrate lyase and malate synthase (GlcB) catalyze the net incorporation of carbon during growth of microorganisms on ac ... >> More

  Establishment or maintenance of a persistent infection by Mycobacterium tuberculosis requires the glyoxylate pathway. This is a bypass of the tricarboxylic acid cycle in which isocitrate lyase and malate synthase (GlcB) catalyze the net incorporation of carbon during growth of microorganisms on acetate or fatty acids as the primary carbon source. The glcB gene from M. tuberculosis, which encodes malate synthase, was cloned, and GlcB was expressed in Escherichia coli. The influence of media conditions on expression in M. tuberculosis indicated that this enzyme is regulated differentially to isocitrate lyase. Purified GlcB had K(m) values of 57 and 30 microm for its substrates glyoxylate and acetyl coenzyme A, respectively, and was inhibited by bromopyruvate, oxalate, and phosphoenolpyruvate. The GlcB structure was solved to 2.1-A resolution in the presence of glyoxylate and magnesium. We also report the structure of GlcB in complex with the products of the reaction, coenzyme A and malate, solved to 2.7-A resolution. Coenzyme A binds in a bent conformation, and the details of its interactions are described, together with implications on the enzyme mechanism. << Less

  J. Biol. Chem. 278:1735-1743(2003) [PubMed] [EuropePMC]

• Purification and properties of malate synthetase.

  DIXON G.H., KORNBERG H.L., LUND P.

  Biochim Biophys Acta 41:217-233(1960) [PubMed] [EuropePMC]