Abstract
The ongoing pandemic with the emergence of immune evasion potential and particularly the current omicron sub variants intensified the situation further. Although vaccine are available but the immune evasion capabilities of the recent variants demand further efficient therapeutic choices to control the SARS-CoV-2 pandemic. Hence, considering the necessity of the small molecule inhibitor we target the main protease (3CLpro) which is an appealing target for the development of anti-viral drugs against the SARS-CoV-2. High-throughput molecular in silico screening of South African natural compounds database (SANCDB) reported as Isojacareubin and Glabranin the potential inhibitors for main protease. The calculated docking scores were reported to be -8.47 kcal/mol and -8.03 kcal/mol respectively. Moreover, the structural-dynamic assessment reported that Isojacareubin in complex with 3CLpro exhibit more stable dynamic behaviour than Glabranin. Inhibition assay indicated that Isojacareubin could inhibit SARS-CoV-2 3CLpro in a time- and dose-dependent manner, with IC50 values of 16.00±1.35 μM (60-min incubation). Next, the covalent binding sites of Isojacareubin on SARS-CoV-2 3CLpro was identified by biomass spectrometry which reported that Isojacareubin can covalently bind to thiols or Cysteine through Michael addition. To evaluate the inactivation potency of Isojacareubin, the inactivation kinetics was further investigated. The inactivation kinetic curves were plotted according to various concentrations with gradient-ascending incubation times. The K I value of Isojacareubin was determined as 30.71 μM, while the K inact value was calculated as 0.054 min-1. These results suggest that Isojacareubin is a covalent inhibitor of SARS-CoV-2 3CLpro.
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