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Identification of potential Leptospira phosphoheptose isomerase inhibitors through virtual high-throughput screening.

ABSTRACT: The life-threatening infections caused by Leptospira serovars demand the need for designing anti-leptospirosis drugs. The present study encompasses exploring inhibitors against phosphoheptose isomerase (GmhA) of Leptospira, which is vital for lipopolysaccharide (LPS) biosynthesis and is identified as a common drug target through the subtractive genomic approach. GmhA model was built in Modeller 9v7. Structural refinement and energy minimization of the predicted model was carried out using Maestro 9.0. The refined model reliability was assessed through Procheck, ProSA, ProQ and Profile 3D. The substrate-based virtual high-throughput screening (VHTS) in Ligand. Info Meta-Database tool generated an in-house library of 354 substrate structural analogs. Furthermore, structure-based VHTS from the in-house library with different conformations of each ligand provided 14 novel competitive inhibitors. The model together with insight gained from the VHTS would be a promising starting point for developing anti-leptospirosis competitive inhibitors targeting LPS biosynthesis pathway.

SUBMITTER: Umamaheswari A

PROVIDER: S-EPMC5054147 | biostudies-literature | 2010 Dec

REPOSITORIES: biostudies-literature

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Identification of potential Leptospira phosphoheptose isomerase inhibitors through virtual high-throughput screening.

Umamaheswari Amineni A Pradhan Dibyabhaba D Hemanthkumar Marisetty M

Genomics, proteomics & bioinformatics 20101201 4

The life-threatening infections caused by Leptospira serovars demand the need for designing anti-leptospirosis drugs. The present study encompasses exploring inhibitors against phosphoheptose isomerase (GmhA) of Leptospira, which is vital for lipopolysaccharide (LPS) biosynthesis and is identified as a common drug target through the subtractive genomic approach. GmhA model was built in Modeller 9v7. Structural refinement and energy minimization of the predicted model was carried out using Maestr ...[more]

PMID: 21382593

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Project description:UNLABELLED:Influenza A virus (IAV) infections cause major morbidity and mortality, generating an urgent need for novel antiviral therapeutics. We recently established a dual myxovirus high-throughput screening protocol that combines a fully replication-competent IAV-WSN strain and a respiratory syncytial virus reporter strain for the simultaneous identification of IAV-specific, paramyxovirus-specific, and broad-spectrum inhibitors. In the present study, this protocol was applied to a screening campaign to assess a diverse chemical library with over 142,000 entries. Focusing on IAV-specific hits, we obtained a hit rate of 0.03% after cytotoxicity testing and counterscreening. Three chemically distinct hit classes with nanomolar potency and favorable cytotoxicity profiles were selected. Time-of-addition, minigenome, and viral entry studies demonstrated that these classes block hemagglutinin (HA)-mediated membrane fusion. Antiviral activity extends to an isolate from the 2009 pandemic and, in one case, another group 1 subtype. Target identification through biolayer interferometry confirmed binding of all hit compounds to HA. Resistance profiling revealed two distinct escape mechanisms: primary resistance, associated with reduced compound binding, and secondary resistance, associated with unaltered binding. Secondary resistance was mediated, unusually, through two different pairs of cooperative mutations, each combining a mutation eliminating the membrane-proximal stalk N-glycan with a membrane-distal change in HA1 or HA2. Chemical synthesis of an analog library combined with in silico docking extracted a docking pose for the hit classes. Chemical interrogation spotlights IAV HA as a major druggable target for small-molecule inhibition. Our study identifies novel chemical scaffolds with high developmental potential, outlines diverse routes of IAV escape from entry inhibition, and establishes a path toward structure-aided lead development. IMPORTANCE:This study is one of the first to apply a fully replication-competent third-generation IAV reporter strain to a large-scale high-throughput screen (HTS) drug discovery campaign, allowing multicycle infection and screening in physiologically relevant human respiratory cells. A large number of potential druggable targets was thus chemically interrogated, but mechanistic characterization, positive target identification, and resistance profiling demonstrated that three chemically promising and structurally distinct hit classes selected for further analysis all block HA-mediated membrane fusion. Viral escape from inhibition could be achieved through primary and secondary resistance mechanisms. In silico docking predicted compound binding to a microdomain located at the membrane-distal site of the prefusion HA stalk that was also previously suggested as a target site for chemically unrelated HA inhibitors. This study identifies an unexpected chemodominance of the HA stalk microdomain for small-molecule inhibitors in IAV inhibitor screening campaigns and highlights a novel mechanism of cooperative resistance to IAV entry blockers.

Dataset Information

Identification of potential Leptospira phosphoheptose isomerase inhibitors through virtual high-throughput screening.

Publications

Identification of potential Leptospira phosphoheptose isomerase inhibitors through virtual high-throughput screening.

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