The Crucial Role of Trehalose and Structurally Related Oligosaccharides in the Biosynthesis and Transfer of Mycolic Acids in Corynebacterineae
Résumé
Glycosylated p-hydroxybenzoic acid methyl esters and structurally related phenolphthiocerol glycolipids are important virulence factors of Mycobacterium tuberculosis. Although both types of molecules are thought to be derived from p-hydroxybenzoic acid, the origin of this putative biosynthetic precursor in mycobacteria remained to be established. We describe the characterization of a transposon mutant of M. tuberculosis deficient in the production of all forms of p-hydroxybenzoic acid derivatives. The transposon was found to be inserted in Rv2949c, a gene located in the vicinity of the polyketide synthase gene pks15/1, involved in the elongation of p-hydroxybenzoate to phenolphthiocerol in phenolic glycolipidproducing strains. A recombinant form of the Rv2949c enzyme was produced in the fast-growing non-pathogenic Mycobacterium smegmatis and purified to near homogeneity. The recombinant enzyme catalyzed the removal of the pyruvyl moiety of chorismate to form p-hydroxybenzoate with an apparent K m value for chorismate of 19.7 M and a k cat value of 0.102 s ؊1. Strong inhibition of the reaction by p-hydroxybenzoate but not by pyruvate was observed. These results establish Rv2949c as a chorismate pyruvate-lyase responsible for the direct conversion of chorismate to p-hydroxybenzoate and identify Rv2949c as the sole enzymatic source of p-hydroxybenzoic acid in M. tuberculosis. The chorismate pathway, present only in bacteria, fungi, and plants, provides a wealth of compounds with diverse biological functions, including aromatic amino acids, folate cofactors, menaquinones, ubiquinones, pigments, and iron-chelating siderophores. Mycobacterium tuberculosis, the etiological agent of tuberculosis in humans, is no exception to the rule, and chorismate in this species is the probable common precursor for the biosynthesis of a series of products (see Fig. 1) that are important both in the physiology and in the pathogenicity of the bacterium. In addition to folate and aromatic amino acids (phenylalanine, tyrosine, and tryptophan), M. tuberculosis produces salicylic acid-derived siderophores known as mycobactins (1), isoprenoid quino
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