GENOMIC ENZYMOLOGY--THE ENOLASE SUPERFAMILY

University Of Illinois Urbana-Champaign, Champaign IL

Investigators

Linked publications & trials

Abstract

DESCRIPTION: The members of the enolase superfamily catalyze different overall reactions that share a common partial reaction in which a stabilized enolate anion is generated by abstraction of the alpha-proton of a carboxylic acid. The enolate anion intermediates are partitioned to different products in the different active sites. The catalytic groups in the structurally characterized members of the superfamily are found in (beta/alpha)-barrel domains. The principal investigator and his group describe structural and mechanistic studies of new members of the superfamily that will provide an understanding of the general strategy used to catalyze the common partial reaction and the subsequent divergent partial reactions to generate different products: 1) D-Glucarate dehydratase from Escherichia coli (GlucD) will be studied so that they can understand how the structure of the barrel domain can be modified to evolve an active site that is homologous to mandelate racemate but additionally catalyzes a dehydration reaction. 2) D-Galactonate dehydratase (GalD) and L-rhamnonate dehydratase (RhamD), both from E. coli, catalyze dehydration reactions on acid sugar substrates that differ in the stereochemistry of the leaving OH group. Dr. Gerlt and his group will study these so that they can determine how Nature varied the structure of the barrel domain to allow the stereochemical courses of this dehydration reaction to differ. 3) Orthologous o-succinylbenzoate synthases (OSBS's) are related by a low degree of sequence identity. The OSBS from Amycolaptosis also catalyzes the racemization of N-acylamino acids. They will explore the hypothesis that the sequence divergence is possible because the substrate is chemically unstable. They will improve the racemase activity of the OSBS from Amycolaptosis, thereby mimicking the process that Nature used to evolve new enzymes from old enzymes. 4) Although carboxyphosphonoenolpyruvate synthase from Streptomyces hygroscopicus (CPEPS) is homologous to enolase, its primary sequence lacks the essential general acid catalyst in the enolase. CPEPS is reported to catalyze a transesterification reaction. Dr. Gerlt and his group will investigate the mechanism of this reaction so that they can better understand the underlying catalytic strategy used by the members of the superfamily.