HUMAN PROINSULIN AND ANALOGS THEREOF AND METHOD OF PREPARATION BY MICROBIAL POLYPEPTIDE EXPRESSION AND CONVERSION THEREOF TO HUMAN INSULIN

摘要

Microbial expression of a chimeric gene is used to produce a polypeptide comprising the amino acid sequence of human proinsulin, or an analog thereof differing in the "C" chain portion. A polypeptide so produced contains a sequence of additional amino acid units sufficient in number to protect it from bacterial proteases, and has a cleavage site e.g. a methionine residue adjacent the sequence of amino acid units corresponding to the proinsulin or proinsulin analog. Cleavage at this site (e.g. by CNBr) generates proinsulin (or the analog) which is treated in vitro to form the disulfide bonds between the "A" and "B" chain proteins characteristic of human insulin. The "C" chain portion is then excised enzymatically to yield human insulin useful e.g. in the treatment of diabetes. The chimeric gene may be synthesised from oligonucleotides and inserted into a plasmid which is used to transform a host cell, e.g. E. coli.