Expression of a New Recombinant Collagenase Protein of Lucilia Sericata in SF9 Insect Cell as a Potential Method for Wound Healing

Iranian Journal of Biotechnology
Hamzeh AlipourSedigheh Zakeri

Abstract

Today, the use of maggot therapy has become widespread due to the increase in chronic ulcers in the world. The recombinant production of secreted enzymes from these larvae is a novel non-invasive method for the treatment of chronic ulcers. Lucilia Sericata (L. sericata) collagenase (MMP-1) has been expressed in insect cells. Collagenase is an enzyme that is widely used in clinical therapy and industry. It has been indicated that collagenase is expressed and secreted in salivary glands of L. sericata while using for maggot debridement therapy. In the present study we decided to produce the recombinant form of collagenase enzyme in Spodoptera frugiperda (SF9) insect cells using the baculovirus expression system (Bac-to-Bac). cloned the coding sequences (residues 494-1705) of L. sericata collagenase into the pFastBacHTA as donor plasmid. After transposition in the bacmid of DH10Bac host, the bacmid was transfected into the Sf9 cell line, then the expressed recombinant collagenase (MMP-1) was purified using the Ni-NTA agarose. The recombinant protein was verified by Western blotting. Furthermore, the biological activity of purified protein was measured in the presence of its specific substrate and its inhibitor, which was 67 IU.mL-...Continue Reading

Datasets Mentioned

BETA
KY612612

Methods Mentioned

BETA
transfection
PCR
Assay
collagenase assay
collagenase

Software Mentioned

Oligo
Oligo7
Gene Runner

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