Characterization of MenA (isoprenyl diphosphate:1,4-dihydroxy-2-naphthoate isoprenyltransferase) from Mycobacterium tuberculosis
| Autor: | Melanie Ikeh, Dean C. Crick, Rakesh K. Dhiman, Venugopal Pujari, James M. Kincaid, Tanya Parish |
|---|---|
| Jazyk: | angličtina |
| Rok vydání: | 2019 |
| Předmět: |
0301 basic medicine
Science Naphthols macromolecular substances 01 natural sciences Chemical synthesis Substrate Specificity Divalent Mycobacterium tuberculosis 03 medical and health sciences Bacterial Proteins Escherichia coli chemistry.chemical_classification Alkyl and Aryl Transferases Microbial Viability Multidisciplinary biology 010405 organic chemistry Genetic Complementation Test Substrate (chemistry) biology.organism_classification Terpenoid 0104 chemical sciences Cytosol 030104 developmental biology Enzyme chemistry Biochemistry Medicine Specific activity Gene Deletion |
| Zdroj: | PLoS ONE, Vol 14, Iss 4, p e0214958 (2019) |
| ISSN: | 1932-6203 |
| Popis: | 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. |
| Databáze: | OpenAIRE |
| Externí odkaz: |
|