Preparation of beta-trypsin by affinity chromatography of enterokinase-activated bovine trypsinogen |
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Authors: | J J Liepnieks A Light |
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Affiliation: | Department of Chemistry, Purdue University, W. Lafayette, Indiana 47907, USA |
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Abstract: | The preparation of β-trypsin can be simply accomplished from the activation of bovine trypsinogen with partially purified enterokinase in the presence of STI-Sepharose. Enterokinase catalyzes the specific cleavage of lysine 6-isoleucine 7 and the presence of STI prevents autolysis of β-trypsin by forming a stable inactive complex. The STI immobilized to Sepharose is suitable for the subsequent purification of the activation mixture by affinity chromatography. Inert protein and contaminants are removed with a buffer at pH 4.5. A change to a buffer at pH 2.6 or the introduction of a pH gradient leads to the recovery of highly purified β-trypsin. The procedure produces β-trypsin in a 70–75% yield, which is essentially a theoretical recovery, and all operations can be completed within 6 hr. |
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