The crystal structure of phenylpyruvate decarboxylase from Azospirillum brasilense at 1.5 Å resolution Implications for its catalytic and regulatory mechanism

• Published in: The FEBS journal Vol. 274; no. 9; pp. 2363 - 2375
• Main Authors: Versées, Wim, Spaepen, Stijn, Vanderleyden, Jos, Steyaert, Jan
• Format: Journal Article
• Language: English
• Published: England 01.05.2007
• Subjects: Allosteric Site - genetics; Amino Acid Substitution - genetics; Azospirillum brasilense - enzymology; Azospirillum brasilense - genetics; Carboxy-Lyases - chemistry; Carboxy-Lyases - genetics; Carboxy-Lyases - metabolism; Catalysis; Cloning, Molecular; Crystallization; Crystallography, X-Ray; Dimerization; Enzyme Activation - genetics; Glutamic Acid - genetics; Leucine - genetics; Models, Chemical; Models, Molecular; Substrate Specificity - genetics
• ISSN: 1742-464X; 1742-4658
• DOI: 10.1111/j.1742-4658.2007.05771.x

Phenylpyruvate decarboxylase (PPDC) of Azospirillum brasilense , involved in the biosynthesis of the plant hormone indole‐3‐acetic acid and the antimicrobial compound phenylacetic acid, is a thiamine diphosphate‐dependent enzyme that catalyses the nonoxidative decarboxylation of indole‐ and phenylpyruvate. Analogous to yeast pyruvate decarboxylases, PPDC is subject to allosteric substrate activation, showing sigmoidal v versus [S] plots. The present paper reports the crystal structure of this enzyme determined at 1.5 Å resolution. The subunit architecture of PPDC is characteristic for other members of the pyruvate oxidase family, with each subunit consisting of three domains with an open α/β topology. An active site loop, bearing the catalytic residues His112 and His113, could not be modelled due to flexibility. The biological tetramer is best described as an asymmetric dimer of dimers. A cysteine residue that has been suggested as the site for regulatory substrate binding in yeast pyruvate decarboxylase is not conserved, requiring a different mechanism for allosteric substrate activation in PPDC. Only minor changes occur in the interactions with the cofactors, thiamine diphosphate and Mg 2+ , compared to pyruvate decarboxylase. A greater diversity is observed in the substrate binding pocket accounting for the difference in substrate specificity. Moreover, a catalytically important glutamate residue conserved in nearly all decarboxylases is replaced by a leucine in PPDC. The consequences of these differences in terms of the catalytic and regulatory mechanism of PPDC are discussed.