GENE FRAGMENT POLYMERIZATION GIVES INCREASED YIELDS OF RECOMBINANT HUMAN PROINSULIN C-PEPTIDE

A multimerization strategy to improve yields upon recombinant producti on of the 31-aa human proinsulin C-peptide is presented. Gene fragment s encoding the C-peptide were assembled using specific head-to-tail mu ltimerization. DNA constructs encoding one, three or seven copies of t he C-peptide gene, fused to a serum albumin binding affinity tag, were expressed intracellularly in Escherichia coli. The three fusion prote ins were produced at similar levels (approximately 50 mg/l) and were p roteolytically stable during production. Enzymatic digestion by trypsi n-carboxypeptidase B treatment of the fusion proteins was shown to eff iciently release native C-peptide, as determined by mass spectrometry, reverse-phase chromatography and a radioimmunoassay. The quantitative yields of C-peptide obtained from the three different fusion proteins suggest that this multimerization strategy could provide a cost-effic ient production scheme for the C-peptide, and that this strategy could be useful also for production of other recombinant peptides. (C) 1998 Elsevier Science B.V.