Construction, high-density fermentation and purification of recombinant enterokinase catalytic subunit in Pichia pastoris

Enterokinase was a serine protein hydroltyic enzyme that could recognize amino acid sequence DDDDK and cleave the peptide bond after "K". cDNA of bovine enterokinase light chain was artificially synthesized based on the sequences in the GeneBank, and inserted into the pPICZαA after validation of obtaining plasmid pPICZαA/EK L, which was transmitted into Pichia pastoris SMD 1168H after linearization, thus the recombinant pichia pastois strain was obtained by high-density fermentation of recombinant pichia pastoris strain, rEK L (recombinant enterokinase light chain, rEK L) purification, N-terminus sequences of EK examination and bioactivity assay. The results demonstrated that rEK L was highly expressed in the culture supernatant with high specificity and the amino acid sequences in N-terminus were consistent with the literature report. Fig 5, Ref 17