Rv0216, a conserved hypothetical protein from Mycobacterium tuberculosis that is essential for bacterial survival during infection, has a double hotdog fold

Abstract

The Mycobacterium tuberculosis genome contains about 4000 genes, of which approximately a third code for proteins of unknown function or are classified as conserved hypothetical proteins. We have determined the three-dimensional structure of one of these, the rv0216 gene product, which has been shown to be essential for M. tuberculosis growth in vivo. The structure exhibits the greatest similarity to bacterial and eukaryotic hydratases that catalyse the R-specific hydration of 2-enoyl coenzyme A. However, only part of the catalytic machinery is conserved in Rv0216 and it showed no activity for the substrate crotonyl-CoA. The structure of Rv0216 allows us to assign new functional annotations to a family of seven other M. tuberculosis proteins, a number if which are essential for bacterial survival during infection and growth.

Heterologous Expression of Mycobacterial Proteins in Saccharomyces cerevisiae Reveals Two Physiologically Functional 3-Hydroxyacyl-Thioester Dehydratases, HtdX and HtdY, in Addition to HadABC and HtdZ

“…Rv0098 is thought to be one of only 219 essential core genes in mycobacteria (33). As an aside, Rv0216, whose gene product was also tested here for complementation, is also a core gene (11). Furthermore, Rv0098 is required for the survival of M. tuberculosis in mouse lung macrophages (40), and has homologues in M. leprae, M. bovis, and M. smegmatis (41).…”

“…The strains used were yPLM35 and yPLM43. (11,41). Of these, Rv0130, denoted HtdZ (21), and Rv0635-Rv0636 and Rv0636-Rv0637, referred to as HadAB and HadBC (9, 37), have since been identified as physiological dehydratases.…”

“…However, several other candidate proteins with putative hydratase-like structures have been suggested as potential HADs, including Rv0098, Rv0216, Rv0241c, Rv0504c, Rv2499c, Rv2524c, Rv3389c, Rv3538, and Rv3542c (11, 41). Within the spectrum of FASII enzymes, dehydratases (along with enoyl reductases) present the highest degree of structural and sequence divergence (11,29,37,38). The classical paradigm is embodied by the Escherichia coli FabA and FabZ proteins, both of which adopt a single "hotdog fold" typically found in dehydratases and thioesterases (13).…”

“…In M. tuberculosis FASII, this step is undertaken by Rv0635-Rv0636 (HadAB) and Rv0636-Rv0637 (HadBC) (9,37) and possibly also Rv0130 (HtdZ) (21,29). However, several other candidate proteins with putative hydratase-like structures have been suggested as potential HADs, including Rv0098, Rv0216, Rv0241c, Rv0504c, Rv2499c, Rv2524c, Rv3389c, Rv3538, and Rv3542c (11,41). Within the spectrum of FASII enzymes, dehydratases (along with enoyl reductases) present the highest degree of structural and sequence divergence (11,29,37,38).…”

Flavonoid inhibitors as novel antimycobacterial agents targeting Rv0636, a putative dehydratase enzyme involved in Mycobacterium tuberculosis fatty acid synthase II

“…Historically, M. tuberculosis FAS-II has proven as a clinically relevant target (Banerjee et al, 1994;Dover et al, 2007;Kremer et al, 2000). Previous analyses by Castell et al (2005) first suggested that Rv0636 of M. tuberculosis H37Rv is a good candidate for the unidentified b-hydroxyacyl-ACP dehydratase of FAS-II. The gene encoding Rv0636 is not only predicted to be essential (Sassetti et al, 2003), but is highly conserved over numerous mycobacterial species such as M. bovis, M. smegmatis and importantly M. leprae.…”