1d8c Citations

Crystal structure of Escherichia coli malate synthase G complexed with magnesium and glyoxylate at 2.0 A resolution: mechanistic implications.

Biochemistry 39 3156-68 (2000)
Cited: 60 times
EuropePMC logo PMID: 10715138

Abstract

The crystal structure of selenomethionine-substituted malate synthase G, an 81 kDa monomeric enzyme from Escherichia coli has been determined by MAD phasing, model building, and crystallographic refinement to a resolution of 2.0 A. The crystallographic R factor is 0.177 for 49 242 reflections observed at the incident wavelength of 1.008 A, and the model stereochemistry is satisfactory. The basic fold of the enzyme is that of a beta8/alpha8 (TIM) barrel. The barrel is centrally located, with an N-terminal alpha-helical domain flanking one side. An inserted beta-sheet domain folds against the opposite side of the barrel, and an alpha-helical C-terminal domain forms a plug which caps the active site. Malate synthase catalyzes the condensation of glyoxylate and acetyl-coenzyme A and hydrolysis of the intermediate to yield malate and coenzyme A, requiring Mg(2+). The structure reveals an enzyme-substrate complex with glyoxylate and Mg(2+) which coordinates the aldehyde and carboxylate functions of the substrate. Two strictly conserved residues, Asp631 and Arg338, are proposed to provide concerted acid-base chemistry for the generation of the enol(ate) intermediate of acetyl-coenzyme A, while main-chain hydrogen bonds and bound Mg(2+) polarize glyoxylate in preparation for nucleophilic attack. The catalytic strategy of malate synthase appears to be essentially the same as that of citrate synthase, with the electrophile activated for nucleophilic attack by nearby positive charges and hydrogen bonds, while concerted acid-base catalysis accomplishes the abstraction of a proton from the methyl group of acetyl-coenzyme A. An active site aspartate is, however, the only common feature of these two enzymes, and the active sites of these enzymes are produced by quite different protein folds. Interesting similarities in the overall folds and modes of substrate recognition are discussed in comparisons of malate synthase with pyruvate kinase and pyruvate phosphate dikinase.

Articles - 1d8c mentioned but not cited (2)



Reviews citing this publication (7)

  1. Indirect use of deuterium in solution NMR studies of protein structure and hydrogen bonding. Tugarinov V. Prog Nucl Magn Reson Spectrosc 77 49-68 (2014)
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Articles citing this publication (51)

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  32. Estimates of methyl 13C and 1H CSA values (Deltasigma) in proteins from cross-correlated spin relaxation. Tugarinov V, Scheurer C, Brüschweiler R, Kay LE. J. Biomol. NMR 30 397-406 (2004)
  33. Selective 1H- 13C NMR spectroscopy of methyl groups in residually protonated samples of large proteins. Guo C, Tugarinov V. J. Biomol. NMR 46 127-133 (2010)
  34. Structure and mechanism of HpcH: a metal ion dependent class II aldolase from the homoprotocatechuate degradation pathway of Escherichia coli. Rea D, Fülöp V, Bugg TD, Roper DI. J. Mol. Biol. 373 866-876 (2007)
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  37. The glcB locus of Rhizobium leguminosarum VF39 encodes an arabinose-inducible malate synthase. García-de los Santos A, Morales A, Baldomá L, Clark SR, Brom S, Yost CK, Hernández-Lucas I, Aguilar J, Hynes MF. Can. J. Microbiol. 48 922-932 (2002)
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  49. Estimating side-chain order in methyl-protonated, perdeuterated proteins via multiple-quantum relaxation violated coherence transfer NMR spectroscopy. Sun H, Godoy-Ruiz R, Tugarinov V. J. Biomol. NMR 52 233-243 (2012)
  50. Simultaneous measurement of ¹H-¹⁵N and methyl ¹Hm-¹³Cm residual dipolar couplings in large proteins. Liao X, Godoy-Ruiz R, Guo C, Tugarinov V. J. Biomol. NMR 51 191-198 (2011)
  51. Dissociation of Mg(ii) and Zn(ii) complexes of simple 2-oxocarboxylates - relationship to CO2 fixation, and the Grignard and Barbier reactions. Miller GBS, Uggerud E. Org. Biomol. Chem. 15 6813-6825 (2017)