Expression and purification of dominant negative protein kinase C isoforms using MBP fusion expression system

초록

Protein kinase C (PKC) isoforms are a family of serine/threonine kinases divided into three subfamilies: classical, novel & atypical, and are important modifiers of several cardiovascular phenomena, carcinogenesis & apoptosis and also promising therapeutic target for cancer and other indications. In our present study, we expressed and purified dominant negative mutants of PKC isoforms. We used pMBPN3 vector to construct expression vectors. The expression vectors were transformed into BL21(DE3) E.coli strain and expressed by IPTG induction. Proteins were expressed as MBP fusion protein and purified by amylase resin. Expression levels were checked by SDS-PAGE analysis. We will use these proteins to check interaction with various PKC interacting proteins to identify whether kinase activity is required for interaction or not.