fumarates and benzylsuccinic-acid

Benzylsuccinate synthase catalyzes a highly unusual reaction: the addition of toluene to fumarate to form (R)-benzylsuccinic acid. The stereochemistry of this reaction has been examined using [d3-methyl]toluene and either fumarate or its cis stereoisomer, maleate, as the substrates. We demonstrate that when fumarate is the cosubstrate, deuterium is transferred from toluene to the C-3 pro-(R) position of benzylsuccinate, implying a syn addition of toluene to the double bond of fumarate. However, when maleate is the cosubstrate, the addition of toluene occurs in an anti fashion, so that deuterium transfer to the C-3 pro-(R) position of benzylsuccinate is also observed. This is consistent with the formation of the C-3 radical of benzylsuccinate as an intermediate, in which rotation about the C-2-C-3 bond can occur to relieve the sterically unfavorable cis conformation of the carboxylate groups when maleate is the cosubstrate.

Topics: Carbon-Carbon Lyases; Deuterium; Fumarates; Hydrogen; Maleates; Nuclear Magnetic Resonance, Biomolecular; Stereoisomerism; Succinates; Toluene

Two dead-end metabolites of anaerobic toluene transformation, benzylsuccinic acid and benzylfumaric acid, accumulated in sulfate-reducing enrichment cultures that were fed toluene as the sole carbon source. Stable isotope-labeled toluene and gas chromatography-mass spectrometry were used to confirm that the compounds resulted from toluene metabolism. The two metabolites constituted less than 10% of the toluene carbon (over 80% was mineralized to carbon dioxide, according to a previous study). This study demonstrates that the novel nonproductive pathway proposed by Evans and coworkers (P. J. Evans, W. Ling, B. Goldschmidt, E. R. Ritter, and L. Y. Young, Appl. Environ. Microbiol. 58:496-501, 1992) for a denitrifying pure culture applies to disparate anaerobic bacteria.

Topics: Bacteria, Anaerobic; Biodegradation, Environmental; Culture Media; Fumarates; Oxidation-Reduction; Succinates; Sulfates; Toluene