Expression of metalloproteinase inhibitors of Hemiscorpius lepturus scorpion venom in bacterial host

Background and Aim : One of the most dangerous endemic scorpions of Iran is Hemiscorpius lepturus that its venom is very toxic and known as responsible of most of scorpion sting dependent deaths in Iran. There are four Metalloproteinase inhibitors in the venom gland transcriptome of this scorpion. Metalloproteinase inhibitors are involved in some of the critical cells activity including the stability and surviving of ECM. This study aimed to investigate the structure and functions of a pair of these molecules and subsequently their cloning and expression for the first time.Methods : bioinformatics analysis and molecular docking on sequences of HLMetInhibit۱ and HLMetInhibit۲ performed (with the MG۷۶۴۵۴۱ and MG۷۶۴۵۴۲ NCBI accession numbers). The recombinant plasmids (pET-۲۲b- HLMetInhibit ۱,۲) designed and synthesized, then transformed into the E. coli BL۲۱ bacterial host. The colony-PCR and electrophoresis on one percent agarose gel performed and protein expression induced by different concentrations of IPTG at different times. Then SDS-PAGE and staining by coomassie brilliant blue and western blotting performed.Results : The highest binding affinity predicted for HLMetInhibit۱ by molecular docking results. Gene cloning performed. Highest protein expression detected after ۵ hours and with ۱ mM concentration of IPTG. SDS-PAGE and western blotting confirmed the protein expression correctness for HLMetInhibit۱ and HLMetInhibit۲ with ۲۶ and ۱۷ KD of molecular weights.Conclusion : Gene cloning and protein expression of Hemiscorpius lepturus metalloproteinase inhibitors۱ and ۲ as the novel proteins that have not been studied so far, is approachable and it is hoped that after further in vitro and in vivo investigations, they will be the suitable candidates for research in the Various therapeutic fields.

کلیدواژه ها:

Cloning and Expression ، Hemiscorpius lepturus ، Metalloproteinase inhibitors