Purification and biochemical characterization of a recombinant mouse seminal vesicle trypsin inhibitor produced in Escherichia coli.

Escherichia coli cells were transformed with an expression vector constructed by inserting a DNA fragment encoding a Kazal-type trypsin inhibitor from mouse seminal vesicle into pGEX-2. The cloned cells were able to produce a high yield of a chimeric polypeptide made by fusing the trypsin inhibitor to glutathione S-transferase. The chimeric polypeptide could be purified through an affinity column of glutathione agarose beads. The purified protein could be digested with thrombin to release the recombinant trypsin inhibitor which could be further purified by HPLC of the thrombin digests on a reverse-phase C4 column. The recombinant trypsin inhibitor was homogeneous and showed trypsin inhibitor activity as strong as that of the naturally occurring trypsin inhibitor.