The Role of a FAD Cofactor in the Regulation of Acetohydroxyacid Synthase by Redox Signaling Molecules

Thierry Lonhienne; Mario D. Garcia; Luke W. Guddat

doi:10.1074/jbc.M116.773242

The Role of a FAD Cofactor in the Regulation of Acetohydroxyacid Synthase by Redox Signaling Molecules*

From the School of Chemistry and Molecular Biosciences, The University of Queensland, Brisbane, 4072 Queensland, Australia

↵1 To whom correspondence may be addressed. E-mail: t.lonhienne{at}uq.edu.au.
↵2 To whom correspondence may be addressed: University of Queensland, Department of Bioscience, School of Chemistry and Molecular Biology, Bldg. 76, Cooper Rd., St. Lucia, QLD, 4072, Australia. Tel.: 61-7-3365-3549; Fax: 61-7-3365-4273; E-mail: luke.guddat{at}uq.edu.au.

Edited by F. Peter Guengerich

Abstract

Acetohydroxyacid synthase (AHAS) catalyzes the first step of branched-chain amino acid (BCAA) biosynthesis, a pathway essential to the lifecycle of plants and microorganisms. This enzyme is of high interest because its inhibition is at the base of the exceptional potency of herbicides and potentially a target for the discovery of new antimicrobial drugs. The enzyme has conserved attributes from its predicted ancestor, pyruvate oxidase, such as a ubiquinone-binding site and the requirement for FAD as cofactor. Here, we show that these requirements are linked to the regulation of AHAS, in relationship to its anabolic function. Using various soluble quinone derivatives (e.g. ubiquinones), we reveal a new path of down-regulation of AHAS activity involving inhibition by oxidized redox-signaling molecules. The inhibition process relies on two factors specific to AHAS: (i) the requirement of a reduced FAD cofactor for the enzyme to be active and (ii) a characteristic slow rate of FAD reduction by the pyruvate oxidase side reaction of the enzyme. The mechanism of inhibition involves the oxidation of the FAD cofactor, leading to a time-dependent inhibition of AHAS correlated with the slow process of FAD re-reduction. The existence and conservation of such a complex mechanism suggests that the redox level of the environment regulates the BCAA biosynthesis pathway. This mode of regulation appears to be the foundation of the inhibitory activity of many of the commercial herbicides that target AHAS.

Footnotes

↵* The work was supported by Grant 1008736 from the National Health and Medical Research Council. The authors declare that they have no conflicts of interest with the contents of this article.