Cloning and Expression of GranzymeM cDNA

Defence against virally infected and tumor cells depends on the action of cytotoxic T lymphocytes and natural killer cells. GranzymeM is a member of a family of granule serine proteases, which is mainly expressed by these cells. GranzymeM appears to be a potent inducer of tumor cell death with morphological hallmarks that are unique among all granzymes.Since the cloning of the granzymeM cDNA in the early 1990s, it has remained an orphan granzyme for many years and only during the past few years the interest in this protease has increased. Therefore in this study The cDNA of activehGzmM, was cloned into bacterial expression vector pET21a(+).for protein expression, Recombinant plasmid was transformed into E. coli BL21(DE3) cells . Recombinant GranzymeM protein was induced at different conditions. Induction condition was optimized to obtain high yield of recombinant proteinand protein expression analyzed by SDS-PAGE. Results showed that the recombinant enzyme has optimum expression at 22°C with 1mM IPTG during 5h and expressed as inclusion bodies in E. coli

Keywords:

Cloning , Tumor cell , Serine protease , GranzymeM , Inclusion body