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Biochemical characterization and chemical validation of Leishmania MAP Kinase-3 as a potential drug target.

ABSTRACT: Protozoan parasites of the Leishmania genus have evolved unique signaling pathways that can sense various environmental changes and trigger stage differentiation for survival and host infectivity. MAP kinase (MAPK) plays a critical role in various cellular activities like cell differentiation, proliferation, stress regulation, and apoptosis. The Leishmania donovani MAPK3 (LdMAPK3) is involved in the regulation of flagella length and hence plays an important role in disease transmission. Here, we reported the gene cloning, protein expression, biochemical characterizations, inhibition studies and cell proliferation assay of LdMAPK3. The recombinant purified LdMAPK3 enzyme obeys the Michaelis-Menten equation with Km and Vmax of LdMAPK3 was found to be 20.23?nM and 38.77?±?0.71 nmoles ATP consumed/mg LdMAPK3/min respectively. The maximum kinase activity of LdMAPK3 was recorded at 35?°C and pH 7. The in-vitro inhibition studies with two natural inhibitors genistein (GEN) and chrysin (CHY) was evaluated against LdMAPK3. The Ki value for GEN and CHY were found to be 3.76?±?0.28?µM and Ki?=?8.75?±?0.11?µM respectively. The IC50 value for the compounds, GEN and CHY against L. donovani promastigotes were calculated as 9.9?µg/mL and 13?µg/mL respectively. Our study, therefore, reports LdMAPK3 as a new target for therapeutic approach against leishmaniasis.

SUBMITTER: Raj S

PROVIDER: S-EPMC6838069 | biostudies-literature | 2019 Nov

REPOSITORIES: biostudies-literature

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Biochemical characterization and chemical validation of Leishmania MAP Kinase-3 as a potential drug target.

Raj Shweta S Saha Gundappa G Sasidharan Santanu S Dubey Vikash Kumar VK Saudagar Prakash P

Scientific reports 20191107 1

Protozoan parasites of the Leishmania genus have evolved unique signaling pathways that can sense various environmental changes and trigger stage differentiation for survival and host infectivity. MAP kinase (MAPK) plays a critical role in various cellular activities like cell differentiation, proliferation, stress regulation, and apoptosis. The Leishmania donovani MAPK3 (LdMAPK3) is involved in the regulation of flagella length and hence plays an important role in disease transmission. Here, we ...[more]

PMID: 31700105

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Project description:BackgroundThe 5'-methylthioadenosine phosphorylase (MTAP), an enzyme involved in purine and polyamine metabolism and in the methionine salvage pathway, is considered as a potential drug target against cancer and trypanosomiasis. In fact, Trypanosoma and Leishmania parasites lack de novo purine pathways and rely on purine salvage pathways to meet their requirements. Herein, we propose the first comprehensive bioinformatic and structural characterization of the putative Leishmania infantum MTAP (LiMTAP), using a comparative computational approach.ResultsSequence analysis showed that LiMTAP shared higher identity rates with the Trypanosoma brucei (TbMTAP) and the human (huMTAP) homologs as compared to the human purine nucleoside phosphorylase (huPNP). Motifs search using MEME identified more common patterns and higher relatedness of the parasite proteins to the huMTAP than to the huPNP. The 3D structures of LiMTAP and TbMTAP were predicted by homology modeling and compared to the crystal structure of the huMTAP. These models presented conserved secondary structures compared to the huMTAP, with a similar topology corresponding to the Rossmann fold. This confirmed that both LiMTAP and TbMTAP are members of the NP-I family. In comparison to the huMTAP, the 3D model of LiMTAP showed an additional α-helix, at the C terminal extremity. One peptide located in this specific region was used to generate a specific antibody to LiMTAP. In comparison with the active site (AS) of huMTAP, the parasite ASs presented significant differences in the shape and the electrostatic potentials (EPs). Molecular docking of 5'-methylthioadenosine (MTA) and 5'-hydroxyethylthio-adenosine (HETA) on the ASs on the three proteins predicted differential binding modes and interactions when comparing the parasite proteins to the human orthologue.ConclusionsThis study highlighted significant structural peculiarities, corresponding to functionally relevant sequence divergence in LiMTAP, making of it a potential drug target against Leishmania.

Dataset Information

Biochemical characterization and chemical validation of Leishmania MAP Kinase-3 as a potential drug target.

Publications

Biochemical characterization and chemical validation of Leishmania MAP Kinase-3 as a potential drug target.

Similar Datasets

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