title: Intraorganellar calcium and pH control proinsulin cleavage in the pancreatic beta cell via two distinct site-specific endopeptidases.
abstract: Insulin is produced from an inactive precursor, proinsulin, through initial endoproteolytic cleavage at sites marked by pairs of basic amino-acid residues. We report here that lysates of insulin secretory granules contain two distinct Ca-dependent acidic endoproteases; one (type I) cleaving exclusively on the C-terminal side of Arg 31.Arg 32 (B-chain/C-peptide junction), the other (type II) preferentially on the C-terminal side of Lys 64.Arg 65 of proinsulin (C-peptide/A-chain junction). The Ca and pH requirements of these proteinases suggested that the type-II proteinase would be active in the Golgi apparatus and the secretory granule, whereas type-I activity would be compatible only with the intragranular environment. Kinetic analyses of (pro)insulin conversion intermediates in [35S]methionine-pulsed rat islets support this supposition. Our results suggest a simple mechanism whereby different dibasic sites can be cleaved in different cellular compartments. In conjunction with the regulation of the ionic composition of such compartments and the operation of post-Golgi segregation, our results also suggest how proteolytic conversion of diverse proproteins destined for different cellular sites can occur differentially and in a regulated manner.
authors.affiliation: Department of Clinical Biochemistry, University of Cambridge, UK.
authors.forename: H W
authors.initials: HW
authors.lastname: Davidson
authors.forename: C J
authors.initials: CJ
authors.lastname: Rhodes
authors.forename: J C
authors.initials: JC
authors.lastname: Hutton
mesh_terms: D000818:Animals; D002118:Calcium / Q000502:physiology*; D003594:Cytoplasmic Granules; D010450:Endopeptidases / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D066298:In Vitro Techniques; D007515:Islets of Langerhans / Q000378:metabolism*; D007700:Kinetics; D011384:Proinsulin / Q000235:genetics*; D011499:Protein Processing, Post-Translational; D051381:Rats
chemical_list: D011384:Proinsulin; D010450:Endopeptidases; C054196:proinsulin endopeptidase I; C061496:proinsulin endopeptidase II; D002118:Calcium