Production of human adrenocorticotropin by cleavage of alkaline‐phosphatase‐derived fusion proteins containing repetitive recognition sequences for collagenases

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Abstract
Recombinant plasmids coding for fusion proteins which consist of human adrenocorticotropin joined to N-terminal sequences of Escherichia coli alkaline phosphatase via collegenase-sensitive linkers were constructed and used for the production of these proteins by transformed E. coli cells. It was shown that repetitive linkers of the form -Gly-(Pro-Xaa-Gly)n-Pro- with n .gtoreq. 2 were cleaved by clostridopeptidase A (Clostridium histolyticum) by orders of magnitude faster than corresponding nonrepetitive sequences (n = 1). The C-terminal cleavage product was Gly-Pro-adrenocorticotropin which could be converted to the authentic hormone by dipeptidyl peptidase IV. On the basis of these enzymatic reactions a procedure for the preparation of pure adrenocorticotropin was developed. Derivatives of alkaline phosphatase containing similar repetitive linker sequences were cleaved by clostridiopeptidase A as efficiently as the adrenocorticotropin fusion proteins.