title: Specific binding of proinsulin C-peptide to human cell membranes.
abstract: Recent reports have demonstrated beneficial effects of proinsulin C-peptide in the diabetic state, including improvements of kidney and nerve function. To examine the background to these effects, C-peptide binding to cell membranes has been studied by using fluorescence correlation spectroscopy. Measurements of ligand-membrane interactions at single-molecule detection sensitivity in 0.2-fl confocal volume elements show specific binding of fluorescently labeled C-peptide to several human cell types. Full saturation of the C-peptide binding to the cell surface is obtained at low nanomolar concentrations. Scatchard analysis of binding to renal tubular cells indicates the existence of a high-affinity binding process with K(ass) > 3.3 x 10(9) M(-1). Addition of excess unlabeled C-peptide is accompanied by competitive displacement, yielding a dissociation rate constant of 4.5 x 10(-4) s(-1). The C-terminal pentapeptide also displaces C-peptide bound to cell membranes, indicating that the binding occurs at this segment of the ligand. Nonnative D-C-peptide and a randomly scrambled C-peptide do not compete for binding with the labeled C-peptide, nor were crossreactions observed with insulin, insulin-like growth factor (IGF)-I, IGF-II, or proinsulin. Pretreatment of cells with pertussis toxin, known to modify receptor-coupled G proteins, abolishes the binding. It is concluded that C-peptide binds to specific G protein-coupled receptors on human cell membranes, thus providing a molecular basis for its biological effects.
authors.affiliation: Department of Medical Biochemistry, Karolinska Institutet, SE-171 77 Stockholm, Sweden.
authors.forename: R
authors.initials: R
authors.lastname: Rigler
authors.forename: A
authors.initials: A
authors.lastname: Pramanik
authors.forename: P
authors.initials: P
authors.lastname: Jonasson
authors.forename: G
authors.initials: G
authors.lastname: Kratz
authors.forename: O T
authors.initials: OT
authors.lastname: Jansson
authors.forename: P
authors.initials: P
authors.lastname: Nygren
authors.forename: S
authors.initials: S
authors.lastname: Stâhl
authors.forename: K
authors.initials: K
authors.lastname: Ekberg
authors.forename: B
authors.initials: B
authors.lastname: Johansson
authors.forename: S
authors.initials: S
authors.lastname: Uhlén
authors.forename: M
authors.initials: M
authors.lastname: Uhlén
authors.forename: H
authors.initials: H
authors.lastname: Jörnvall
authors.forename: J
authors.initials: J
authors.lastname: Wahren
mesh_terms: D001667:Binding, Competitive; D002096:C-Peptide / Q000378:metabolism*; D002462:Cell Membrane / Q000187:drug effects / Q000378:metabolism; D006801:Humans; D007684:Kidney Tubules / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D007700:Kinetics; D008856:Microscopy, Fluorescence; D037342:Pertussis Toxin; D010566:Virulence Factors, Bordetella / Q000494:pharmacology
chemical_list: D002096:C-Peptide; D010566:Virulence Factors, Bordetella; D037342:Pertussis Toxin