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Mesaconyl-Coenzyme A Hydratase, a New Enzyme of Two Central Carbon Metabolic Pathways in Bacteria

Jan Zarzycki,¹ Ansgar Schlichting,¹ Nina Strychalsky,¹ Michael Müller,² Birgit E. Alber,^1, and Georg Fuchs^1*

Mikrobiologie, Fakultät für Biologie, Universität Freiburg, Freiburg, Germany,¹ Pharmazeutische und Medizinische Chemie, Fakultät für Chemie, Pharmazie und Geowissenschaften, Universität Freiburg, Freiburg, Germany²

Received 5 October 2007/ Accepted 28 November 2007

The coenzyme A (CoA)-activated C₅-dicarboxylic acids mesaconyl-CoA and β-methylmalyl-CoA play roles in two as yet not completely resolved central carbon metabolic pathways in bacteria. First, these compounds are intermediates in the 3-hydroxypropionate cycle for autotrophic CO₂ fixation in Chloroflexus aurantiacus, a phototrophic green nonsulfur bacterium. Second, mesaconyl-CoA and β-methylmalyl-CoA are intermediates in the ethylmalonyl-CoA pathway for acetate assimilation in various bacteria, e.g., in Rhodobacter sphaeroides, Methylobacterium extorquens, and Streptomyces species. In both cases, mesaconyl-CoA hydratase was postulated to catalyze the interconversion of mesaconyl-CoA and β-methylmalyl-CoA. The putative genes coding for this enzyme in C. aurantiacus and R. sphaeroides were cloned and heterologously expressed in Escherichia coli, and the proteins were purified and studied. The recombinant homodimeric 80-kDa proteins catalyzed the reversible dehydration of erythro-β-methylmalyl-CoA to mesaconyl-CoA with rates of 1,300 µmol min^–1 mg protein^–1. Genes coding for similar enzymes with two (R)-enoyl-CoA hydratase domains are present in the genomes of Roseiflexus, Methylobacterium, Hyphomonas, Rhodospirillum, Xanthobacter, Caulobacter, Magnetospirillum, Jannaschia, Sagittula, Parvibaculum, Stappia, Oceanicola, Loktanella, Silicibacter, Roseobacter, Roseovarius, Dinoroseobacter, Sulfitobacter, Paracoccus, and Ralstonia species. A similar yet distinct class of enzymes containing only one hydratase domain was found in various other bacteria, such as Streptomyces species. The role of this widely distributed new enzyme is discussed.

Published ahead of print on 7 December 2007.

Present address: Department of Microbiology, The Ohio State University, 484 West 12th Ave., Columbus, OH 43210-1292.