Mycobacterium tuberculosis is a bacterium responsible for causing approximately 10.4 million tuberculosis cases around the world each year. As tuberculosis incidence, drug resistance, and prevalence rates stay high, new antibiotics are in great need and demand. The rv3802c gene is a member of the five-gene cluster starting from rv3799c to rv3804c, a cluster involved in mycolic acid biosynthesis. While enzymatic roles for most of this cluster’s enzymes have been elucidated, less is certain about the rv3802c gene product beyond its thioesterase and phospholipase activity. Two expression constructs were designed and tested, and resulting purification steps yielded highly pure enzyme. Development of a preliminary assay aided in inhibition studies, with the fluorometric assay using resorufin butyrate as the substrate. This work culminated in the successful elucidation of the first M. tuberculosis Rv3802 X-ray crystal structure to a 1.7 Å resolution. Comparisons between the Rv3802 structure and the structure of its apo homolog, Mycobacterium smegmatis (MSMEG_6394), revealed interesting differences. Ultimately, this research provides a basis for drug discovery efforts targeting Rv3802, as an expansive, hydrophobic active site has been revealed. Additionally, this work provides structural insights to a potential biological function.

Purification, crystallization, and characterization of essential Mycobacterium tuberculosis Rv3802

255th National Meeting (2018)

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