l-Asparaginase as a new molecular target against leishmaniasis: insights into the mechanism of action and structure-based inhibitor design

Abstract

l-Asparaginases belong to a family of amidohydrolases that catalyze the conversion of l-asparagine into l-aspartic acid and ammonia. Although bacterial l-asparaginases have been used extensively as anti-leukemic agents, their possible role as potential drug targets for pathogenic organisms has not been explored. The presence of genes coding for putative l-asparaginase enzymes in the Leishmania donovani genome hinted towards the specific role of these enzymes in extending survival benefit to the organism. To investigate whether this enzyme can serve as a potential drug target against the Leishmania pathogen, we obtained structural models of one of the putative Leishmania l-asparaginase I (LdAI). Using an integrated computational approach involving molecular modelling, docking and molecular dynamics simulations, we found crucial differences between catalytic residues of LdAI as compared to bacterial l-asparaginases. The deviation from the canonical acid-base pair at triad I, along with the structural reorganization of a β-hairpin loop in the presence of a substrate, indicated an altogether new mechanism of action of the LdAI enzyme. Moreover, the finding of compositional and functional differences between LdAI and human asparaginase was used as a criterion to identify specific small molecule inhibitors. Through virtual screening of a library of 11 438 compounds, we report five compounds that showed favorable interactions with the active pocket of LdAI, without adversely affecting human asparaginase. One of these compounds when tested on cultured Leishmania promastigotes displayed a promising leishmanicidal effect. Overall, our work not only provides first hand mechanistic insights of LdAI but also proposes five strongly active compounds which may prove as effective anti-leishmaniasis molecules.

Identification and validation of l‐asparaginase as a potential metabolic target against Mycobacterium tuberculosis

“…Therefore, no phagolysosomal complex is formed and pathogen easily replicates inside the host macrophages. Similar conditions have been observed in case of Leishmania donovani , and inhibitors against its l ‐asparaginase were successful in their intended purpose …”

“…Redocking of selected compounds from all the libraries (filtered through ADMET) with MtA and human l ‐asparaginase (HLA) was performed by Molecular Operating Environment v2009.10 (MOE) . Details of active site grid parameterization and free energy calculations are described elsewhere …”

L-Asparaginase of Leishmania donovani: Metabolic target and its role in Amphotericin B resistance

“…However, enthalpy contribution in binding of L2 was higher than L1. This could be due to higher H-bond interactions of L2 with seven residues of the active pocket namely Thr 38, Leu 49, His 192, Asp 85 and Ser 87, Asp 118 and Met 144 (Singh et al., 2015). However, L1 interactions mediated through single H-bond with Thr 38 and five other H-bonds with four residues viz.…”

“…We further characterized LdAI structurally and functionally. Earlier we had modeled the structure of this enzyme and proposed potential inhibitors, L1 and L2 (Singh et al., 2015). In this study we report that these inhibitors (albeit at high concentrations but non-toxic to humans) could effectively retard the growth of Leishmania indicating necessity of LdAI to the parasite.…”

HAT2 mediates histone H4K4 acetylation and affects micrococcal nuclease sensitivity of chromatin in Leishmania donovani

“…Leishmania donovani Ag83 promastigotes were cultured in M199 media with Hanks salt (Gibco) at pH 7.4 as described previously [35]. It was supplemented with 10% FBS (Gibco) and 100U/ml penicillin streptomycin (Gibco) as described previously.…”