Publications

• The missing piece of the type II fatty acid synthase system from Mycobacterium tuberculosis.

  Sacco E., Covarrubias A.S., O'Hare H.M., Carroll P., Eynard N., Jones T.A., Parish T., Daffe M., Backbro K., Quemard A.

  The Mycobacterium tuberculosis fatty acid synthase type II (FAS-II) system has the unique property of producing unusually long-chain fatty acids involved in the biosynthesis of mycolic acids, key molecules of the tubercle bacillus. The enzyme(s) responsible for dehydration of (3R)-hydroxyacyl-ACP ... >> More

  The Mycobacterium tuberculosis fatty acid synthase type II (FAS-II) system has the unique property of producing unusually long-chain fatty acids involved in the biosynthesis of mycolic acids, key molecules of the tubercle bacillus. The enzyme(s) responsible for dehydration of (3R)-hydroxyacyl-ACP during the elongation cycles of the mycobacterial FAS-II remained unknown. This step is classically catalyzed by FabZ- and FabA-type enzymes in bacteria, but no such proteins are present in mycobacteria. Bioinformatic analyses and an essentiality study allowed the identification of a candidate protein cluster, Rv0635-Rv0636-Rv0637. Its expression in recombinant Escherichia coli strains leads to the formation of two heterodimers, Rv0635-Rv0636 (HadAB) and Rv0636-Rv0637 (HadBC), which also occurs in Mycobacterium smegmatis, as shown by split-Trp assays. Both heterodimers exhibit the enzymatic properties expected for mycobacterial FAS-II dehydratases: a marked specificity for both long-chain (>or=C(12)) and ACP-linked substrates. Furthermore, they function as 3-hydroxyacyl dehydratases when coupled with MabA and InhA enzymes from the M. tuberculosis FAS-II system. HadAB and HadBC are the long-sought (3R)-hydroxyacyl-ACP dehydratases. The correlation between the substrate specificities of these enzymes, the organization of the orthologous gene cluster in different Corynebacterineae, and the structure of their mycolic acids suggests distinct roles for both heterodimers during the elongation process. This work describes bacterial monofunctional (3R)-hydroxyacyl-ACP dehydratases belonging to the hydratase 2 family. Their original structure and the fact that they are essential for M. tuberculosis survival make these enzymes very good candidates for the development of antimycobacterial drugs. << Less

  Proc Natl Acad Sci U S A 104:14628-14633(2007) [PubMed] [EuropePMC]

  This publication is cited by 3 other entries.

• Rv3389C from Mycobacterium tuberculosis, a member of the (R)-specific hydratase/dehydratase family.

  Sacco E., Legendre V., Laval F., Zerbib D., Montrozier H., Eynard N., Guilhot C., Daffe M., Quemard A.

  The (R)-specific 3-hydroxyacyl dehydratases/trans-enoyl hydratases are key proteins in the biosynthesis of fatty acids. In mycobacteria, such enzymes remain unknown, although they are involved in the biosynthesis of major and essential lipids like mycolic acids. First bioinformatic analyses allowe ... >> More

  The (R)-specific 3-hydroxyacyl dehydratases/trans-enoyl hydratases are key proteins in the biosynthesis of fatty acids. In mycobacteria, such enzymes remain unknown, although they are involved in the biosynthesis of major and essential lipids like mycolic acids. First bioinformatic analyses allowed to identify a single candidate protein, namely Rv3389c, that belongs to the hydratases 2 family and is most likely made of a distinctive asymmetric double hot dog fold. The purified recombinant Rv3389c protein was shown to efficiently catalyze the hydration of (C(8)-C(16)) enoyl-CoA substrates. Furthermore, it catalyzed the dehydration of a 3-hydroxyacyl-CoA in coupled reactions with both reductases (MabA and InhA) of the acyl carrier protein (ACP)-dependent M. tuberculosis fatty acid synthase type II involved in mycolic acid biosynthesis. Yet, the facts that Rv3389c activity decreased in the presence of ACP, versus CoA, derivative and that Rv3389c knockout mutant had no visible variation of its fatty acid content suggested the occurrence of additional hydratase/dehydratase candidates. Accordingly, further and detailed bioinformatic analyses led to the identification of other members of the hydratases 2 family in M. tuberculosis. << Less

  Biochim. Biophys. Acta 1774:303-311(2007) [PubMed] [EuropePMC]

  This publication is cited by 3 other entries.

• Revisiting the assignment of Rv0241c to fatty acid synthase type II of Mycobacterium tuberculosis.

  Sacco E., Slama N., Baeckbro K., Parish T., Laval F., Daffe M., Eynard N., Quemard A.

  The fatty acid synthase type II enzymatic complex of Mycobacterium tuberculosis (FAS-II(Mt)) catalyzes an essential metabolic pathway involved in the biosynthesis of major envelope lipids, mycolic acids. The partner proteins of this singular FAS-II system represent relevant targets for antitubercu ... >> More

  The fatty acid synthase type II enzymatic complex of Mycobacterium tuberculosis (FAS-II(Mt)) catalyzes an essential metabolic pathway involved in the biosynthesis of major envelope lipids, mycolic acids. The partner proteins of this singular FAS-II system represent relevant targets for antituberculous drug design. Two heterodimers of the hydratase 2 protein family, HadAB and HadBC, were shown to be involved in the (3R)-hydroxyacyl-ACP dehydration (HAD) step of FAS-II(Mt) cycles. Recently, an additional member of this family, Rv0241c, was proposed to have the same function, based on the heterologous complementation of a HAD mutant of the yeast mitochondrial FAS-II system. In the present work, Rv0241c was able to complement a HAD mutant in the Escherichia coli model but not a dehydratase-isomerase deficient mutant. However, an enzymatic study of the purified protein demonstrated that Rv0241c possesses a broad chain length specificity for the substrate, unlike FAS-II(Mt) enzymes. Most importantly, Rv0241c exhibited a strict dependence on the coenzyme A (CoA) as opposed to AcpM, the natural acyl carrier protein bearing the chains elongated by FAS-II(Mt). The deletion of Rv0241c showed that this gene is not essential to M. tuberculosis survival in vitro. The resulting mutant did not display any change in the mycolic acid profile. This demonstrates that Rv0241c is a trans-2-enoyl-CoA hydratase/3-hydroxyacyl-CoA dehydratase that does not belong to FAS-II(Mt). The relevance of a heterologous complementation strategy to identifying proteins of such a system is questioned. << Less

  J. Bacteriol. 192:4037-4044(2010) [PubMed] [EuropePMC]

  This publication is cited by 4 other entries.