title: Proinsulin C-peptide activates α-enolase: implications for C-peptide--cell membrane interaction.
abstract: Proinsulin C-peptide shows beneficial effects on microvascular complications of Type 1 diabetes. However, the possible occurrence of membrane C-peptide receptor(s) has not been elucidated. The aim of this study was to identify and characterize membrane proteins to which C-peptide binds. The enzyme α-enolase was co-immunoprecipitated with C-peptide after chemical cross-linking to HL-60 cell surface proteins and identified by mass spectrometry. Recombinant α-enolase activity was modulated by C-peptide, with a significant decrease in K(m) for 2-phosphoglycerate without affecting V(max). The enzyme modulation by C-peptide was abolished when C-terminal basic lysine residue (K434) of the enzyme was replaced by neutral alanine or acidic glutamate, but not with basic arginine. The enzyme modulation by C-peptide was reproduced with the C-peptide fragments containing glutamate corresponding to position 27 (E27) of the full-length C-peptide. Addition of a lysine analogue to the assay and A31 cell culture abrogated the enzyme modulation and MAP kinase activation by C-peptide, respectively. The results indicate that C-peptide has the capacity to activate α-enolase through a specific interaction between E27 of the peptide and K434 of the enzyme. Since α-enolase plays a role as a cell surface receptor for plasminogen, it may conceivably also serve as a receptor for C-peptide in vivo.
authors.affiliation: Department of Biomedical Sciences, Graduate School of Veterinary Medicine, Hokkaido University, Sapporo 060-0818, Japan.
authors.forename: Tatsuya
authors.initials: T
authors.lastname: Ishii
authors.forename: Keigo
authors.initials: K
authors.lastname: Fukano
authors.forename: Kohei
authors.initials: K
authors.lastname: Shimada
authors.forename: Akihiro
authors.initials: A
authors.lastname: Kamikawa
authors.forename: Yuko
authors.initials: Y
authors.lastname: Okamatsu-Ogura
authors.forename: Akira
authors.initials: A
authors.lastname: Terao
authors.forename: Toshihide
authors.initials: T
authors.lastname: Yoshida
authors.forename: Masayuki
authors.initials: M
authors.lastname: Saito
authors.forename: Kazuhiro
authors.initials: K
authors.lastname: Kimura
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D002096:C-Peptide / Q000378:metabolism*; D002462:Cell Membrane / Q000378:metabolism*; D002478:Cells, Cultured; D018922:HL-60 Cells; D006801:Humans; D008969:Molecular Sequence Data; D010751:Phosphopyruvate Hydratase / Q000378:metabolism*
chemical_list: D002096:C-Peptide; D010751:Phosphopyruvate Hydratase