Publications

• Role of betaine:CoA ligase (CaiC) in the activation of betaines and the transfer of coenzyme A in Escherichia coli.

  Bernal V., Arense P., Blatz V., Mandrand-Berthelot M.A., Canovas M., Iborra J.L.

  <h4>Aims</h4>Characterization of the role of CaiC in the biotransformation of trimethylammonium compounds into l(-)-carnitine in Escherichia coli.<h4>Methods and results</h4>The caiC gene was cloned and overexpressed in E. coli and its effect on the production of l(-)-carnitine was analysed. Betai ... >> More

  <h4>Aims</h4>Characterization of the role of CaiC in the biotransformation of trimethylammonium compounds into l(-)-carnitine in Escherichia coli.<h4>Methods and results</h4>The caiC gene was cloned and overexpressed in E. coli and its effect on the production of l(-)-carnitine was analysed. Betaine:CoA ligase and CoA transferase activities were analysed in cell free extracts and products were studied by electrospray mass spectrometry (ESI-MS). Substrate specificity of the caiC gene product was high, reflecting the high specialization of the carnitine pathway. Although CoA-transferase activity was also detected in vitro, the main in vivo role of CaiC was found to be the synthesis of betainyl-CoAs. Overexpression of CaiC allowed the biotransformation of crotonobetaine to l(-)-carnitine to be enhanced nearly 20-fold, the yield reaching up to 30% (with growing cells). Higher yields were obtained using resting cells (up to 60%), even when d(+)-carnitine was used as substrate.<h4>Conclusions</h4>The expression of CaiC is a control step in the biotransformation of trimethylammonium compounds in E. coli.<h4>Significance and impact of the study</h4>A bacterial betaine:CoA ligase has been characterized for the first time, underlining its important role for the production of l-carnitine with Escherichia coli. << Less

  J. Appl. Microbiol. 105:42-50(2008) [PubMed] [EuropePMC]

  This publication is cited by 3 other entries.

• Molecular characterization of the cai operon necessary for carnitine metabolism in Escherichia coli.

  Eichler K., Bourgis F., Buchet A., Kleber H.-P., Mandrand-Berthelot M.-A.

  The sequence encompassing the cai genes of Escherichia coli, which encode the carnitine pathway, has been determined. Apart from the already identified caiB gene coding for the carnitine dehydratase, five additional open reading frames were identified. They belong to the caiTABCDE operon, which wa ... >> More

  The sequence encompassing the cai genes of Escherichia coli, which encode the carnitine pathway, has been determined. Apart from the already identified caiB gene coding for the carnitine dehydratase, five additional open reading frames were identified. They belong to the caiTABCDE operon, which was shown to be located at the first minute on the chromosome and transcribed during anaerobic growth in the presence of carnitine. The activity of carnitine dehydratase was dependent on the CRP regulatory protein and strongly enhanced in the absence of a functional H-NS protein, in relation to the consensus sequences detected in the promoter region of the cai operon. In vivo expression studies led to the synthesis of five polypeptides in addition to CaiB, with predicted molecular masses of 56,613 Da (CaiT), 42,564 Da (CaiA), 59,311 Da (CaiC), 32,329 Da (CaiD) and 21,930 Da (CaiE). Amino acid sequence similarity or enzymatic analysis supported the function assigned to each protein. CaiT was suggested to be the transport system for carnitine or betaines, CaiA an oxidoreduction enzyme, and CaiC a crotonobetaine/carnitine CoA ligase. CaiD bears strong homology with enoyl hydratases/isomerases. Overproduction of CaiE was shown to stimulate the carnitine racemase activity of the CaiD protein and to markedly increase the basal level of carnitine dehydratase activity. It is inferred that CaiE is an enzyme involved in the synthesis or the activation of the still unknown cofactor required for carnitine dehydratase and carnitine racemase activities. Taken together, these data suggest that the carnitine pathway in E. coli resembles that found in a strain situated between Agrobacterium and Rhizobium. << Less

  Mol. Microbiol. 13:775-786(1994) [PubMed] [EuropePMC]