Abstract
BackgroundThe secretory proteins of Mycobacterium tuberculosis (M. tuberculosis) have been known to be involved in the virulence, pathogenesis as well as proliferation of the pathogen. Among this set, many proteins have been hypothesized to play a critical role at the genesis of the onset of infection, the primary site of which is invariably the human lung.Methodology/Principal FindingsDuring our efforts to isolate potential binding partners of key secretory proteins of M. tuberculosis from a human lung protein library, we isolated peptides that strongly bound the virulence determinant protein Esat6. All peptides were less than fifty amino acids in length and the binding was confirmed by in vivo as well as in vitro studies. Curiously, we found all three binders to be unusually rich in phenylalanine, with one of the three peptides a short fragment of the human cytochrome c oxidase-3 (Cox-3). The most accessible of the three binders, named Hcl1, was shown also to bind to the Mycobacterium smegmatis (M. smegmatis) Esat6 homologue. Expression of hcl1 in M. tuberculosis H37Rv led to considerable reduction in growth. Microarray analysis showed that Hcl1 affects a host of key cellular pathways in M. tuberculosis. In a macrophage infection model, the sets expressing hcl1 were shown to clear off M. tuberculosis in much greater numbers than those infected macrophages wherein the M. tuberculosis was not expressing the peptide. Transmission electron microscopy studies of hcl1 expressing M. tuberculosis showed prominent expulsion of cellular material into the matrix, hinting at cell wall damage.Conclusions/SignificanceWhile the debilitating effects of Hcl1 on M. tuberculosis are unrelated and not because of the peptide's binding to Esat6–as the latter is not an essential protein of M. tuberculosis–nonetheless, further studies with this peptide, as well as a closer inspection of the microarray data may shed important light on the suitability of such small phenylalanine-rich peptides as potential drug-like molecules against this pathogen.
Highlights
Tuberculosis (TB), a disease caused by M. tuberculosis, is completely curable, and yet, two million succumb to it every year [1,2]
Isolation of Peptides That Bind Esat6 In order to search for host interacting partners of Esat6, bacterial two-hybrid reporter strain was co-transformed with esat6pBTnn and human lung cDNA library cloned in pTRG
We report isolation and characterization of phenylalanine-rich peptides that bind Esat6 protein of M. tuberculosis
Summary
Tuberculosis (TB), a disease caused by M. tuberculosis, is completely curable, and yet, two million succumb to it every year [1,2]. In India, that along with sub-Saharan Africa has the largest number of TB cases, partial adherence to directly observed drug treatment regimen, coupled with non-availability of the drugs in remote areas combine devastatingly to exacerbate the problem, resulting in multi-drug resistant strains that de facto necessitate the scientific community’s search for newer anti-TB molecules [3,4,5] Tied with this seemingly intractable predicament is the lengthy anti-TB therapy that lasts on average six to eight months, giving ample opportunity for poor patients to play truant [6]. The secretory proteins of Mycobacterium tuberculosis (M. tuberculosis) have been known to be involved in the virulence, pathogenesis as well as proliferation of the pathogen Among this set, many proteins have been hypothesized to play a critical role at the genesis of the onset of infection, the primary site of which is invariably the human lung
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