title: Effect of modification of connecting peptide of proinsulin on its export.
abstract: The production of a high yield of proinsulin using a secretion vector has been difficult, even with such modifications to the vector as a strong promoter and a good ribosome binding site. This investigation was therefore initiated to see whether modification of the connecting peptide of proinsulin has any effect on the export of proinsulin. We constructed three types of proinsulin secretion vectors: (a) pEZZ18-PI, by inserting the proinsulin gene into pEZZ18 vector; (b) pEZZ18-PI-C, by modifying ZZ-proinsulin by addition of the carboxy terminal peptide region of human insulin-like growth factor I (hIGFI) to the carboxy terminal end of proinsulin; and (c) pEZZ18-PI analogues, by sequentially deleting the connecting peptide region of proinsulin. The highest export yield of proinsulin was obtained when the connecting peptide region of the proinsulin was similar in size to that of hIGFI, or when most of the connecting peptide region of the proinsulin was deleted. The amount of exported ZZ-proinsulin analogues in these clones was over 25-times higher than that of ZZ-proinsulin with an unmodified connecting peptide in the secretion/expression vector pEZZ18-PI. On the basis of these observations, we conclude that modification of the mature domain of proinsulin is a critical factor for determination of the export of proinsulin.
authors.affiliation: Department of Microbiology and Infectious Diseases, Faculty of Medicine, University of Calgary, Alberta, Canada.
authors.forename: Y
authors.initials: Y
authors.lastname: Kang
authors.forename: J W
authors.initials: JW
authors.lastname: Yoon
mesh_terms: D000595:Amino Acid Sequence; D001483:Base Sequence; D005822:Genetic Vectors; D007334:Insulin-Like Growth Factor I / Q000096:biosynthesis / Q000235:genetics; D008969:Molecular Sequence Data; D011384:Proinsulin / Q000096:biosynthesis* / Q000235:genetics
chemical_list: D007334:Insulin-Like Growth Factor I; D011384:Proinsulin