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Characterization of MenA (isoprenyl diphosphate:1,4-dihydroxy-2-naphthoate isoprenyltransferase) from Mycobacterium tuberculosis.
Dhiman, Rakesh K; Pujari, Venugopal; Kincaid, James M; Ikeh, Melanie A; Parish, Tanya; Crick, Dean C.
Afiliación
  • Dhiman RK; Department of Microbiology, Immunology and Pathology, Colorado State University, Fort Collins, CO, United States of America.
  • Pujari V; Department of Microbiology, Immunology and Pathology, Colorado State University, Fort Collins, CO, United States of America.
  • Kincaid JM; Department of Microbiology, Immunology and Pathology, Colorado State University, Fort Collins, CO, United States of America.
  • Ikeh MA; Queen Mary University of London, Barts & The London School of Medicine and Dentistry, London, United Kingdom.
  • Parish T; Queen Mary University of London, Barts & The London School of Medicine and Dentistry, London, United Kingdom.
  • Crick DC; TB Discovery Research, Infectious Disease Research Institute, Seattle, WA, United States of America.
PLoS One ; 14(4): e0214958, 2019.
Article en En | MEDLINE | ID: mdl-30978223
The menaquinone biosynthetic pathway presents a promising drug target against Mycobacterium tuberculosis and potentially other Gram-positive pathogens. In the present study, the essentiality, steady state kinetics of MenA from M. tuberculosis and the mechanism of MenA inhibition by Ro 48-8071 were characterized. MenA [isoprenyl diphosphate:1,4-dihydroxy-2-naphthoate (DHNA) isoprenyltransferase] catalyzes a critical reaction in menaquinone biosynthesis that involves the conversion of cytosolic DHNA, to membrane bound demethylmenaquinone by transferring a hydrophobic 45-carbon isoprenoid chain (in the case of mycobacteria) to the ring nucleus of DHNA. Rv0534c previously identified as the gene encoding MenA in M. tuberculosis complemented a menA deletion in E. coli and an E. coli host expressing Rv0534c exhibited an eight-fold increase in MenA specific activity over the control strain harboring empty vector under similar assay conditions. Expression of Rv0534c is essential for mycobacterial survival and the native enzyme from M. tuberculosis H37Rv was characterized using membrane preparations as it was not possible to solubilize and purify the recombinant enzyme. The enzyme is absolutely dependent on the presence of a divalent cation for optimal activity with Mg+2 being the most effective and is active over a wide pH range, with pH 8.5 being optimal. The apparent Km values for DHNA and farnesyl diphosphate were found to be 8.2 and 4.3 µM, respectively. Ro 48-8071, a compound previously reported to inhibit mycobacterial MenA activity, is non-competitive with regard to DHNA and competitive with regard to the isoprenyldiphosphate substrate.
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Texto completo: 1 Colección: 01-internacional Base de datos: MEDLINE Asunto principal: Proteínas Bacterianas / Transferasas Alquil y Aril / Viabilidad Microbiana / Mycobacterium tuberculosis Idioma: En Revista: PLoS One Asunto de la revista: CIENCIA / MEDICINA Año: 2019 Tipo del documento: Article País de afiliación: Estados Unidos Pais de publicación: Estados Unidos
Texto completo
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XML
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Texto completo: 1 Colección: 01-internacional Base de datos: MEDLINE Asunto principal: Proteínas Bacterianas / Transferasas Alquil y Aril / Viabilidad Microbiana / Mycobacterium tuberculosis Idioma: En Revista: PLoS One Asunto de la revista: CIENCIA / MEDICINA Año: 2019 Tipo del documento: Article País de afiliación: Estados Unidos Pais de publicación: Estados Unidos