Publications

• Studies with substrate and cofactor analogues provide evidence for a radical mechanism in the chorismate synthase reaction.

  Osborne A., Thorneley R.N., Abell C., Bornemann S.

  Chorismate synthase catalyzes the conversion of 5-enolpyruvylshikimate 3-phosphate (EPSP) to chorismate. The strict requirement for a reduced FMN cofactor and a trans-1,4-elimination are unusual. (6R)-6-Fluoro-EPSP was shown to be converted to chorismate stoichiometrically with enzyme-active sites ... >> More

  Chorismate synthase catalyzes the conversion of 5-enolpyruvylshikimate 3-phosphate (EPSP) to chorismate. The strict requirement for a reduced FMN cofactor and a trans-1,4-elimination are unusual. (6R)-6-Fluoro-EPSP was shown to be converted to chorismate stoichiometrically with enzyme-active sites in the presence of dithionite. This conversion was associated with the oxidation of FMN to give a stable flavin semiquinone. The IC(50) of the fluorinated substrate analogue was 0.5 and 250 microm with the Escherichia coli enzyme, depending on whether it was preincubated with the enzyme or not. The lack of dissociation of the flavin semiquinone and chorismate from the enzyme appears to be the basis of the essentially irreversible inhibition by this analogue. A dithionite-dependent transient formation of flavin semiquinone during turnover of (6S)-6-fluoro-EPSP has been observed. These reactions are best rationalized by radical chemistry that is strongly supportive of a radical mechanism occurring during normal turnover. The lack of activity with 5-deaza-FMN provides additional evidence for the role of flavin in catalysis by the E. coli enzyme. << Less

  J. Biol. Chem. 275:35825-35830(2000) [PubMed] [EuropePMC]

• Properties of chorismate synthase in Neurospora crassa.

  Gaertner F.H., Cole K.W.

  J. Biol. Chem. 248:4602-4609(1973) [PubMed] [EuropePMC]

• The enzymic synthesis of chorismic and prephenic acids from 3-enolpyruvylshikimic acid 5-phosphate.

  Morell H., Clark M.J., Knowles P.F., Sprinson D.B.

  J Biol Chem 242:82-90(1967) [PubMed] [EuropePMC]

  This publication is cited by 1 other entry.

• A secondary beta deuterium kinetic isotope effect in the chorismate synthase reaction.

  Bornemann S., Theoclitou M.E., Brune M., Webb M.R., Thorneley R.N., Abell C.

  Chorismate synthase (EC 4.6.1.4) is the shikimate pathway enzyme that catalyzes the conversion of 5-enolpyruvylshikimate 3-phosphate (EPSP) to chorismate. The enzyme reaction is unusual because it involves a trans-1,4 elimination of the C-3 phosphate and the C-6 proR hydrogen and it has an absolut ... >> More

  Chorismate synthase (EC 4.6.1.4) is the shikimate pathway enzyme that catalyzes the conversion of 5-enolpyruvylshikimate 3-phosphate (EPSP) to chorismate. The enzyme reaction is unusual because it involves a trans-1,4 elimination of the C-3 phosphate and the C-6 proR hydrogen and it has an absolute requirement for reduced flavin. Several mechanisms have been proposed to account for the cofactor requirement and stereochemistry of the reaction, including a radical mechanism. This paper describes the synthesis of [4-(2)H]EPSP and the observation of kinetic isotope effects using this substrate with both Neurospora crassa and Escherichia coli chorismate synthases. The magnitude of the effects were (D)(V) = 1.08 +/- 0.01 for the N. crassa enzyme and 1.10 +/-0.02 on phosphate release under single-turnover conditions for the E. coli enzyme. The effects are best rationalised as substantial secondary beta isotope effects. It is most likely that the C(3)-O bond is cleaved first in a nonconcerted E1 or radical reaction mechanism. Although this study alone cannot rule out a concerted E2-type mechanism, the C(3)-O bond would have to be substantially more broken than the proR C(6)-H bond in a transition state of such a mechanism. Importantly, although the E. coli and N. crassa enzymes have different rate limiting steps, their catalytic mechanisms are most likely to be chemically identical. Copyright 2000 Academic Press. << Less

  Bioorg. Chem. 28:191-204(2000) [PubMed] [EuropePMC]

• The transient kinetics of Escherichia coli chorismate synthase: substrate consumption, product formation, phosphate dissociation, and characterization of a flavin intermediate.

  Bornemann S., Lowe D.J., Thorneley R.N.

  Chorismate synthase is the seventh enzyme of the shikimate pathway and catalyzes the conversion of 5-enolpyruvylshikimate 3-phosphate (EPSP) to chorismate. The reaction involves the 1,4-elimination of phosphate and the C-(6proR) hydrogen of the substrate with unusual anti stereochemistry and requi ... >> More

  Chorismate synthase is the seventh enzyme of the shikimate pathway and catalyzes the conversion of 5-enolpyruvylshikimate 3-phosphate (EPSP) to chorismate. The reaction involves the 1,4-elimination of phosphate and the C-(6proR) hydrogen of the substrate with unusual anti stereochemistry and requires a reduced flavin cofactor. This paper describes the kinetics of the formation and decay of a flavin intermediate, EPSP consumption, chorismate and phosphate formation, and phosphate dissociation during single and multiple turnover experiments, determined using rapid reaction techniques. The kinetics of phosphate dissociation using the substrate analogues (6R)-[6-2H]EPSP and (6S)-6-fluoro-EPSP have also been determined. The observations are consistent with a nonconcerted chorismate synthase reaction. The flavin intermediate is not simply associated with the conversion of substrate to product because it forms before the substrate is consumed. The transient spectral changes must be associated primarily with events such as protonation of the reduced flavin, a charge transfer complex between reduced flavin and an aromatic amino acid, or a conformational change in the protein. This does not rule out the direct role of flavin in catalysis. << Less

  Biochemistry 35:9907-9916(1996) [PubMed] [EuropePMC]

• Chorismate synthase of Neurospora crassa: a flavoprotein.

  Welch G.R., Cole K.W., Gaertner F.H.

  Arch. Biochem. Biophys. 165:505-518(1974) [PubMed] [EuropePMC]