title: p-Hydroxybenzoic acid synthesis in Mycobacterium tuberculosis.
abstract: 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 glycolipid-producing 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 microm 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.
authors.affiliation: Unité deGénétique Mycobactérienne, Institut Pasteur, 75015, Paris, France.
authors.forename: Gustavo
authors.initials: G
authors.lastname: Stadthagen
authors.forename: Jana
authors.initials: J
authors.lastname: Korduláková
authors.forename: Ruth
authors.initials: R
authors.lastname: Griffin
authors.forename: Patricia
authors.initials: P
authors.lastname: Constant
authors.forename: Iveta
authors.initials: I
authors.lastname: Bottová
authors.forename: Nathalie
authors.initials: N
authors.lastname: Barilone
authors.forename: Brigitte
authors.initials: B
authors.lastname: Gicquel
authors.forename: Mamadou
authors.initials: M
authors.lastname: Daffé
authors.forename: Mary
authors.initials: M
authors.lastname: Jackson
mesh_terms: D002827:Chorismic Acid / Q000378:metabolism; D004251:DNA Transposable Elements / Q000235:genetics; D004926:Escherichia coli / Q000235:genetics; D008745:Methylation; D009154:Mutation; D020102:Mycobacterium smegmatis / Q000235:genetics; D009169:Mycobacterium tuberculosis / Q000201:enzymology* / Q000235:genetics; D007652:Oxo-Acid-Lyases / Q000235:genetics / Q000378:metabolism; D010226:Parabens / Q000737:chemistry / Q000378:metabolism*; D010641:Phenotype; D048630:Polyketide Synthases / Q000235:genetics; D019289:Pyruvic Acid / Q000378:metabolism; D011994:Recombinant Proteins / Q000378:metabolism; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
chemical_list: D004251:DNA Transposable Elements; D010226:Parabens; D011994:Recombinant Proteins; D048630:Polyketide Synthases; D019289:Pyruvic Acid; D007652:Oxo-Acid-Lyases; C075960:chorismate pyruvate lyase; D002827:Chorismic Acid; C038193:4-hydroxybenzoic acid