Exoproteome and Secretome Derived Broad Spectrum Novel Drug and Vaccine Candidates in <em>Vibrio cholerae</em> Targeted by <em>Piper betel</em> Derived Compounds

<div><p><em>Vibrio cholerae</em> is the causal organism of the cholera epidemic, which is mostly prevalent in developing and underdeveloped countries. However, incidences of cholera in developed countries are also alarming. Because of the emergence of new drug-resistant strains, even though several generic drugs and vaccines have been developed over time, <em>Vibrio</em> infections remain a global health problem that appeals for the development of novel drugs and vaccines against the pathogen. Here, applying comparative proteomic and reverse vaccinology approaches to the exoproteome and secretome of the pathogen, we have identified three candidate targets (<em>ompU</em>, <em>uppP</em> and <em>yajC</em>) for most of the pathogenic <em>Vibrio</em> strains. Two targets (<em>uppP</em> and <em>yajC</em>) are novel to <em>Vibrio</em>, and two targets (<em>uppP</em> and <em>ompU</em>) can be used to develop both drugs and vaccines (dual targets) against broad spectrum <em>Vibrio</em> serotypes. Using our novel computational approach, we have identified three peptide vaccine candidates that have high potential to induce both B- and T-cell-mediated immune responses from our identified two dual targets. These two targets were modeled and subjected to virtual screening against natural compounds derived from <em>Piper betel</em>. Seven compounds were identified first time from <em>Piper betel</em> to be highly effective to render the function of these targets to identify them as emerging potential drugs against <em>Vibrio</em>. Our preliminary validation suggests that these identified peptide vaccines and <em>betel</em> compounds are highly effective against <em>Vibrio cholerae</em>. Currently we are exhaustively validating these targets, candidate peptide vaccines, and <em>betel</em> derived lead compounds against a number of <em>Vibrio</em> species.</p> </div

Protein-protein and host-pathogen interactions among ten preliminary identified <i>Vibrio</i> targets.

<p>The interactions demonstrate that the finally selected three targets (<i>ompU</i>, <i>yajC</i>, and <i>uppP</i>) are involved in <i>Vibrio</i> pathogenesis and modulate host response (immunity and apoptosis) by interacting with the host protein PDCD6.</p

The best seven <i>Piper betel</i> compounds that may render activities of <i>Vibrio</i> targets <i>ompU</i> and <i>uppP</i>.

<p>GOLD fitness and Moldock scores were considered to select the compounds. Guineesine, Pinoresinol, and Piperdardine inhibit both targets. Dehydropipernonaline and Piperrolein B are effective on <i>ompU</i>. Chlorogenic acid and Eugenyl acetate are good ligands for <i>uppP</i>.</p

Pepitope analysis of identified T-cell epitopes for their exomembrane topology (colored in red) within the corresponding folded proteins.

<p>A) The “VTSGEPVHS” epitope of <i>uppP</i>, B) the “VTETNAAKY” epitope of <i>ompU</i>, and C) the “YNNAETAKK” epitope of <i>ompU</i>.</p

3D models of <i>Vibrio</i> targets constructed using threading approaches.

<p>A) Front view of <i>ompU</i>, B) Side view of <i>ompU</i>, and C) Front view of <i>uppP</i>.</p

Anti-<i>Vibrio</i> activity of Piperdardine.

<p>A). Growth inhibition effects Piperdardine, Ampicillin, and Chloranphenicol on <i>V. Cholerae O1 Inaba</i> growth as per the disk diffusion method. 1) 100 mM, 2) 200 mM, and 3) 300 mM Piperdardine; 4) water; 5) Ampicillin (10 µg); and 6) Chloranphenicol (30 µg). The zones of inhibition (mm) around disks containing Piperdardine are concentration-dependent: 1) 19.3±0.03; 2) 26.23±0.1; 3) 28.65±0.16. Controls: 4) 0±0; 5) 18.51±0.16; and 6) 29.47±0.16. B). Effects on Piperdardine and Chloranphenicol on <i>V. Cholerae O1 Inaba</i> growth as per the Colony-forming units (CFU/ml) assay. As per the method described in the text, 60 mM of Piperdardine (squares) shows anti-<i>Vibrio</i> effect similar to 100 µg/ml of Chloramphenicol (triangles).</p