Inhibitor binding studies on enoyl reductase reveal conformational changes related to substrate recognition

Anna Roujeinikova, Svetlana Sedelnikova, Gert Jan De Boer, Antoine R. Stuitje, Antoni R. Slabas, John B. Rafferty, David W. Rice

Research output: Contribution to journalArticleResearchpeer-review

Abstract

Enoyl acyl carrier protein reductase (ENR) is involved in fatty acid biosynthesis. In Escherichia coli this enzyme is the target for the experimental family of antibacterial agents, the diazaborines, and for triclosan, a broad spectrum antimicrobial agent. Biochemical studies have suggested that the mechanism of diazaborine inhibition is dependent on NAD+ and not NADH, and resistance of Brassica napus ENR to diazaborines is thought to be due to the replacement of a glycine in the active site of the E. coli enzyme by an alanine at position 138 in the plant homologue. We present here an x-ray analysis of crystals of B. napus ENR A138G grown in the presence of either NAD+ or NADH and the structures of the corresponding ternary complexes with thienodiazaborine obtained either by soaking the drug into the crystals or by co-crystallization of the mutant with NAD+ and diazaborine. Analysis of the ENR A138G complex with diazaborine and NAD+ shows that the site of diazaborine binding is remarkably close to that reported for E. coli ENR. However, the structure of the ternary ENR A138G-NAD+-diazaborine complex obtained using co-crystallization reveals a previously unobserved conformational change affecting 11 residues that flank the active site and move closer to the nicotinamide moiety making extensive van der Waals contacts with diazaborine. Considerations of the mode of substrate binding suggest that this conformational change may reflect a structure of ENR that is important in catalysis.

Original languageEnglish
Pages (from-to)30811-30817
Number of pages7
JournalJournal of Biological Chemistry
Volume274
Issue number43
DOIs
Publication statusPublished - 22 Oct 1999
Externally publishedYes

Cite this

Roujeinikova, A., Sedelnikova, S., De Boer, G. J., Stuitje, A. R., Slabas, A. R., Rafferty, J. B., & Rice, D. W. (1999). Inhibitor binding studies on enoyl reductase reveal conformational changes related to substrate recognition. Journal of Biological Chemistry, 274(43), 30811-30817. https://doi.org/10.1074/jbc.274.43.30811
Roujeinikova, Anna ; Sedelnikova, Svetlana ; De Boer, Gert Jan ; Stuitje, Antoine R. ; Slabas, Antoni R. ; Rafferty, John B. ; Rice, David W. / Inhibitor binding studies on enoyl reductase reveal conformational changes related to substrate recognition. In: Journal of Biological Chemistry. 1999 ; Vol. 274, No. 43. pp. 30811-30817.
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abstract = "Enoyl acyl carrier protein reductase (ENR) is involved in fatty acid biosynthesis. In Escherichia coli this enzyme is the target for the experimental family of antibacterial agents, the diazaborines, and for triclosan, a broad spectrum antimicrobial agent. Biochemical studies have suggested that the mechanism of diazaborine inhibition is dependent on NAD+ and not NADH, and resistance of Brassica napus ENR to diazaborines is thought to be due to the replacement of a glycine in the active site of the E. coli enzyme by an alanine at position 138 in the plant homologue. We present here an x-ray analysis of crystals of B. napus ENR A138G grown in the presence of either NAD+ or NADH and the structures of the corresponding ternary complexes with thienodiazaborine obtained either by soaking the drug into the crystals or by co-crystallization of the mutant with NAD+ and diazaborine. Analysis of the ENR A138G complex with diazaborine and NAD+ shows that the site of diazaborine binding is remarkably close to that reported for E. coli ENR. However, the structure of the ternary ENR A138G-NAD+-diazaborine complex obtained using co-crystallization reveals a previously unobserved conformational change affecting 11 residues that flank the active site and move closer to the nicotinamide moiety making extensive van der Waals contacts with diazaborine. Considerations of the mode of substrate binding suggest that this conformational change may reflect a structure of ENR that is important in catalysis.",
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Inhibitor binding studies on enoyl reductase reveal conformational changes related to substrate recognition. / Roujeinikova, Anna; Sedelnikova, Svetlana; De Boer, Gert Jan; Stuitje, Antoine R.; Slabas, Antoni R.; Rafferty, John B.; Rice, David W.

In: Journal of Biological Chemistry, Vol. 274, No. 43, 22.10.1999, p. 30811-30817.

Research output: Contribution to journalArticleResearchpeer-review

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AU - Roujeinikova, Anna

AU - Sedelnikova, Svetlana

AU - De Boer, Gert Jan

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AU - Slabas, Antoni R.

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AU - Rice, David W.

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AB - Enoyl acyl carrier protein reductase (ENR) is involved in fatty acid biosynthesis. In Escherichia coli this enzyme is the target for the experimental family of antibacterial agents, the diazaborines, and for triclosan, a broad spectrum antimicrobial agent. Biochemical studies have suggested that the mechanism of diazaborine inhibition is dependent on NAD+ and not NADH, and resistance of Brassica napus ENR to diazaborines is thought to be due to the replacement of a glycine in the active site of the E. coli enzyme by an alanine at position 138 in the plant homologue. We present here an x-ray analysis of crystals of B. napus ENR A138G grown in the presence of either NAD+ or NADH and the structures of the corresponding ternary complexes with thienodiazaborine obtained either by soaking the drug into the crystals or by co-crystallization of the mutant with NAD+ and diazaborine. Analysis of the ENR A138G complex with diazaborine and NAD+ shows that the site of diazaborine binding is remarkably close to that reported for E. coli ENR. However, the structure of the ternary ENR A138G-NAD+-diazaborine complex obtained using co-crystallization reveals a previously unobserved conformational change affecting 11 residues that flank the active site and move closer to the nicotinamide moiety making extensive van der Waals contacts with diazaborine. Considerations of the mode of substrate binding suggest that this conformational change may reflect a structure of ENR that is important in catalysis.

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