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Proc Natl Acad Sci U S A. 1987 Sep; 84(17): 6184–6188.
PMCID: PMC299034

Use of a synthetic peptide antigen to generate antisera reactive with a proteolytic processing site in native human proinsulin: demonstration of cleavage within clathrin-coated (pro)secretory vesicles.


Polyclonal antibodies reactive with a cleavage site in human proinsulin (HPI) (C-peptide-A-chain junction) have been raised (rabbit, guinea pig) using a synthetic peptide antigen coupled with keyhole limpet hemocyanin. These antisera recognize native HPI and des-31,32-HPI equally well but react 20-50 times less well with des-64,65-HPI, the intermediate cleaved at the C-peptide-A-chain junction and lacking the Lys-Arg pair. The guinea pig antisera did not recognize insulin but reacted weakly with C peptide at high concentrations; the rabbit antisera reacted with neither insulin nor C peptide. Immunocytochemical studies with human islet tissue localized the immunoreactivity of these antisera to clathrin-coated (pro)secretory vesicles derived from the trans Golgi, indicating that cleavage of the C-peptide-A-chain junction of proinsulin occurs mainly, if not exclusively, in this compartment of the beta cell.

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Selected References

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  • Steiner DF, Docherty K, Carroll R. Golgi/granule processing of peptide hormone and neuropeptide precursors: a minireview. J Cell Biochem. 1984;24(2):121–130. [PubMed]
  • Shibasaki Y, Kawakami T, Kanazawa Y, Akanuma Y, Takaku F. Posttranslational cleavage of proinsulin is blocked by a point mutation in familial hyperproinsulinemia. J Clin Invest. 1985 Jul;76(1):378–380. [PMC free article] [PubMed]
  • Rholam M, Nicolas P, Cohen P. Precursors for peptide hormones share common secondary structures forming features at the proteolytic processing sites. FEBS Lett. 1986 Oct 20;207(1):1–6. [PubMed]
  • Leszczynski JF, Rose GD. Loops in globular proteins: a novel category of secondary structure. Science. 1986 Nov 14;234(4778):849–855. [PubMed]
  • Taussig R, Scheller RH. The Aplysia FMRFamide gene encodes sequences related to mammalian brain peptides. DNA. 1986 Dec;5(6):453–461. [PubMed]
  • Thim L, Hansen MT, Norris K, Hoegh I, Boel E, Forstrom J, Ammerer G, Fiil NP. Secretion and processing of insulin precursors in yeast. Proc Natl Acad Sci U S A. 1986 Sep;83(18):6766–6770. [PMC free article] [PubMed]
  • Steiner DF. The biosynthesis of insulin: genetic, evolutionary, and pathophysiologic aspects. Harvey Lect. 1982 1983;78:191–228. [PubMed]
  • Kemmler W, Peterson JD, Steiner DF. Studies on the conversion of proinsulin to insulin. I. Conversion in vitro with trypsin and carboxypeptidase B. J Biol Chem. 1971 Nov 25;246(22):6786–6791. [PubMed]
  • Houghten RA. General method for the rapid solid-phase synthesis of large numbers of peptides: specificity of antigen-antibody interaction at the level of individual amino acids. Proc Natl Acad Sci U S A. 1985 Aug;82(15):5131–5135. [PMC free article] [PubMed]
  • Walker LE, Ketler TA, Houghten RA, Schulz G, Chersi A, Reisfeld RA. Human major histocompatibility complex class I antigens: residues 61-83 of the HLA-B7 heavy chain specify an alloreactive site. Proc Natl Acad Sci U S A. 1985 Jan;82(2):539–542. [PMC free article] [PubMed]
  • Madsen OD, Frank BH, Steiner DF. Human proinsulin-specific antigenic determinants identified by monoclonal antibodies. Diabetes. 1984 Oct;33(10):1012–1016. [PubMed]
  • Armbruster BL, Carlemalm E, Chiovetti R, Garavito RM, Hobot JA, Kellenberger E, Villiger W. Specimen preparation for electron microscopy using low temperature embedding resins. J Microsc. 1982 Apr;126(Pt 1):77–85. [PubMed]
  • Roth J, Bendayan M, Orci L. Ultrastructural localization of intracellular antigens by the use of protein A-gold complex. J Histochem Cytochem. 1978 Dec;26(12):1074–1081. [PubMed]
  • Cohen RM, Given BD, Licinio-Paixao J, Provow SA, Rue PA, Frank BH, Root MA, Polonsky KS, Tager HS, Rubenstein AH. Proinsulin radioimmunoassay in the evaluation of insulinomas and familial hyperproinsulinemia. Metabolism. 1986 Dec;35(12):1137–1146. [PubMed]
  • Schwartz TW. The processing of peptide precursors. 'Proline-directed arginyl cleavage' and other monobasic processing mechanisms. FEBS Lett. 1986 May 5;200(1):1–10. [PubMed]
  • Welsh M, Hammer RE, Brinster RL, Steiner DF. Stimulation of growth hormone synthesis by glucose in islets of Langerhans isolated from transgenic mice. J Biol Chem. 1986 Oct 5;261(28):12915–12917. [PubMed]
  • Moore HH, Kelly RB. Re-routing of a secretory protein by fusion with human growth hormone sequences. Nature. 1986 May 22;321(6068):443–446. [PubMed]
  • Tramontano A, Janda KD, Lerner RA. Catalytic antibodies. Science. 1986 Dec 19;234(4783):1566–1570. [PubMed]
  • Pollack SJ, Jacobs JW, Schultz PG. Selective chemical catalysis by an antibody. Science. 1986 Dec 19;234(4783):1570–1573. [PubMed]
  • Orci L, Halban P, Amherdt M, Ravazzola M, Vassalli JD, Perrelet A. A clathrin-coated, Golgi-related compartment of the insulin secreting cell accumulates proinsulin in the presence of monensin. Cell. 1984 Nov;39(1):39–47. [PubMed]
  • Orci L, Ravazzola M, Amherdt M, Madsen O, Vassalli JD, Perrelet A. Direct identification of prohormone conversion site in insulin-secreting cells. Cell. 1985 Sep;42(2):671–681. [PubMed]
  • Madsen OD, Cohen RM, Fitch FW, Rubenstein AH, Steiner DF. The production and characterization of monoclonal antibodies specific for human proinsulin using a sensitive microdot assay procedure. Endocrinology. 1983 Dec;113(6):2135–2144. [PubMed]
  • Mojsov S, Heinrich G, Wilson IB, Ravazzola M, Orci L, Habener JF. Preproglucagon gene expression in pancreas and intestine diversifies at the level of post-translational processing. J Biol Chem. 1986 Sep 5;261(25):11880–11889. [PubMed]
  • Orci L, Ravazzola M, Storch MJ, Anderson RG, Vassalli JD, Perrelet A. Proteolytic maturation of insulin is a post-Golgi event which occurs in acidifying clathrin-coated secretory vesicles. Cell. 1987 Jun 19;49(6):865–868. [PubMed]
  • Sando H, Borg J, Steiner DF. Studies on the secretion of newly synthesized proinsulin and insulin from isolated rat islets of Langerhans. J Clin Invest. 1972 Jun;51(6):1476–1485. [PMC free article] [PubMed]

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