DOUBLE-C-PEPTIDE HUMAN PROINSULIN

A fusion gene encoding double-C-peptide human proinsulin was construct ed by insertion of a DNA fragment encoding human C-peptide into the 5' -terminal C-peptide coding sequence of a synthetic human proinsulin ge ne with correct reading frame and over-expressed in E.coli. The purifi ed double-C-peptide human proinsulin shows decreased activity in recep tor binding and insulin immune assays as compared with human proinsuli n. Disulphide bond reconstitution studies demonstrate that there is no t much more influence of the protein concentration on the yield of ref olded double-C-peptide human proinsulin. The double-C-peptide human pr oinsulin shows a 1.86-fold human C-peptide immune activity as compared with that of human proinsulin and gives a good yield of the molecule with correct disulphide bonds in reconstitution studies strongly sugge sting the existence of very flexible conformation of the C-peptide.