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Journal of Bacteriology, August 2006, p. 5479-5486, Vol. 188, No. 15
0021-9193/06/$08.00+0     doi:10.1128/JB.00514-06
Copyright © 2006, American Society for Microbiology. All Rights Reserved.

The FAD-Dependent Tricarballylate Dehydrogenase (TcuA) Enzyme of Salmonella enterica Converts Tricarballylate into cis-Aconitate

Department of Bacteriology, University of Wisconsin—Madison, Madison, Wisconsin 53726

Received 11 April 2006/ Accepted 19 May 2006

Tricarballylate is the causative agent of grass tetany, a ruminant disease characterized by acute magnesium deficiency. Tricarballylate toxicity has been attributed to its ability to chelate magnesium and to inhibit aconitase, a Krebs cycle enzyme. Neither the ruminant nor the normal rumen flora can catabolize tricarballylate to ameliorate its toxic effects. However, the gram-negative enterobacterium Salmonella enterica can use tricarballylate as a carbon and energy source, providing an opportunity to study the genes and enzymes required for tricarballylate catabolism. The tricarballylate utilization (tcu) genes are organized into two transcriptional units, i.e., tcuR and tcuABC. Here, we report the initial biochemical analysis of TcuA. TcuA catalyzed the oxidation of tricarballylate to cis-aconitate. The apparent K_m of TcuA for tricarballylate was 3.8 ± 0.4 mM, with a V_max of 7.9 ± 0.3 mM min^–1, turnover number (k_cat) of 6.7 x 10^–2 s^–1, and a catalytic efficiency (k_cat/K_m) of 17.8 M^–1 s^–1. Optimal activity was measured at pH 7.5 and 30°C. The enzyme was inactivated at 45°C. One mole of FAD was present per mole of TcuA. We propose a role for TcuB as an electron shuttle protein responsible for oxidizing FADH₂ back to FAD in TcuA.

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This article has been cited by other articles:

• Boyd, J. M., Lewis, J. A., Escalante-Semerena, J. C., Downs, D. M. (2008). Salmonella enterica Requires ApbC Function for Growth on Tricarballylate: Evidence of Functional Redundancy between ApbC and IscU. J. Bacteriol. 190: 4596-4602 [Abstract] [Full Text]