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Discovery of novel inhibitors for Leishmania nucleoside diphosphatase kinase (NDK) based on its structural and functional characterization

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Abstract

Nucleoside diphosphate kinases (NDKs) are ubiquitous enzymes that catalyze the transfer of the γ-phosphate moiety from an NTP donor to an NDP acceptor, crucial for maintaining the cellular level of nucleoside triphosphates (NTPs). The inability of trypanosomatids to synthesize purines de novo and their dependence on the salvage pathway makes NDK an attractive target to develop drugs for the diseases they cause. Here we report the discovery of novel inhibitors for Leishmania NDK based on the structural and functional characterization of purified recombinant NDK from Leishmania amazonensis. Recombinant LaNDK possesses auto-phosphorylation, phosphotransferase and kinase activities with Histidine 117 playing an essential role. LaNDK crystals were grown by hanging drop vapour diffusion method in a solution containing 18% PEG-MME 500, 100 mM Bis-Tris propane pH 6.0 and 50 mM MgCl2. It belongs to the hexagonal space group P6322 with unit cell parameters a = b = 115.18, c = 62.18 Å and α = β = 90°, γ = 120°. The structure solved by molecular replacement methods was refined to crystallographic R-factor and Rfree values of 22.54 and 26.52%, respectively. Molecular docking and dynamics simulation-based virtual screening identified putative binding compounds. Protein inhibition studies of selected hits identified five inhibitors effective at micromolar concentrations. One of the compounds showed ~45% inhibition of Leishmania promastigotes proliferation. Analysis of inhibitor-NDK complexes reveals the mode of their binding, facilitating design of new compounds for optimization of activities as drugs against leishmaniasis.

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Acknowledgements

Arjun K. Mishra and Pragati Agnihotri acknowledge fellowship from Indian Council of Medical Research. Nidhi Singh acknowledges fellowship from Council for Scientific and Industrial Research (CSIR). Authors also thank CSIR for funding received under the network project HOPE. Dr. Babu A. Manjasetty, Dr. Hassan Belrhali and ESRF BM14 are duly acknowledged for data collection. BKK and KP Chang were supported by US-NIH Grant # AI-20486. This manuscript bears CDRI communication Number 9320.

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Correspondence to M. I. Siddiqi or J. Venkatesh Pratap.

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Mishra, A.K., Singh, N., Agnihotri, P. et al. Discovery of novel inhibitors for Leishmania nucleoside diphosphatase kinase (NDK) based on its structural and functional characterization. J Comput Aided Mol Des 31, 547–562 (2017). https://doi.org/10.1007/s10822-017-0022-9

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