ISOLA TION AND CHARACTERIZATION OF LEISHMANIAL ADENINE AMINOHYDROLASE AS A DRUG TARGET

Abstract

CUITently available dl:UgS for Leishmaniasis are insufficient in providing promising results. Pentavalent Antimonial does offer first-line drug therapy, however emergence of drug resistant strains have reduced the efficacy of marketed drugs. On the other hand, second-line drug therapy is less effective due to toxicity and high prices, and therefore encouraging new methods of exploring drug targets. Recently discovered enzyme Adenine Aminohydrolase (AHH), counterpart of human purine salvage pathway, is the crucial enzyme that de-aminates Adenine as a substrate in Leishmania spp, which is important for adenine utilization as a purine and a nitrogen source. Characterization in lower species sUPPOlted AHH as a plausible drug target. In the current study, we have confirmed the presence of Adenine Aminohydrolase in given Pakistani Leishmanial strain source. The knowledge of functional partners of targeted enzyme pathway and its mode of kinetic action are important in developing robust enzyme assay and in approximating Kill and Villax values for enzyme identification. Enzyme exhibits Kill value equal to 14.78±5f!M and V III ax value ofO.2098±0.01~lM min/mg demonstrating catalytic efficiency (Kcatl Kill) of 1.27 x 107 M-'sec-'. Insilico- molecular docking software GOLD probed compound [C26H ,sN30 7S;benzyl amino group] i.e. (2-(2-(5-methy(/uran-2yl)-5- nitro-l H-benzo [dJ imidazole-l-ylsu([onyl) anthracene-9, lO-dione with high score calculating sound binding affinity of -6.48 Kcallmole having hydrophobic interactions formed in majority among all the ligand-protein i.e. 2ICS and compound interactions, thus most likely inhibit its activity at non- competitive site. In vitro kinetic analysis estimates ICso value of 0.81 06f!g/ml showing mixed non-competitive inhibition validating molecular docking results.