Soluble expression and secretion of natural N-terminal recombinant chicken cystatin

Objective To carry out the secretive expression of natural N-terminal chicken cystatin(cC).Methods Recombinant expression plasmid p PICZαA-cC was constructed by designing of distinct primers and inserting of cC cDNA into yeast expression vector p PICZαA.The recombinant cC was expressed in Pichia pastoris X-33 after methanol induction.Results SDS-PAGE,Substrate SDS-PAGE and Native-PA GE analysis indicated that after the treatment of β-ME and GuHCl with different combination,the destructive level of disulfide bond in recombinant cC is increased.Treatment by 5mol/L GuHCl and 10%β-ME can break the intra-molecular disulfide bond almost entirely.Conclusion 92.9% of the recombinant cC had intra-molecular disulfide bonds and a similar inhibitory activity against papain with native cC.