Des-(27-31)C-peptide. A novel secretory product of the rat pancreatic beta cell produced by truncation of proinsulin connecting peptide in secretory granules.
Insulin and connecting peptide (C-peptide) are produced in equimolar amounts during proinsulin conversion in the pancreatic beta cell secretory granule. To determine whether insulin and C-peptide are equally stable in beta cell granules (and thus secreted in equimolar amounts), neonatal and adult rat beta cells were pulse-chased, and radiolabeled insulin and C-peptide analyzed by high performance liquid chromatography. A novel truncated C-peptide was identified and shown by mass spectrometry to be des-(27-31)C-peptide (loss of 5 C-terminal amino acids). Des-(27-31)C-peptide is a major beta cell secretory product, accounting for 37.4 +/- 1.6% (neonatal) and 8.5 +/- 0.6% (adult) of total labeled C-peptide in secretory granules after 10 h of chase. Des-(27-31)C-peptide is also secreted in a glucose-sensitive manner from the perfused adult rat pancreas, accounting for approximately 10% of total C-peptide immunoreactivity secreted. Human C-peptide is also a substrate for truncation in granules. Thus, when human proinsulin was expressed (infection with recombinant adenovirus) in transformed (INS) rat beta cells, human des-(27-31)C-peptide was secreted along with the intact human peptide and both intact and truncated rat C-peptide. In addition to truncation, 33.1 +/- 1.2% of C-peptide in neonatal but not adult rat beta cell granules was further degraded. Such degradation was completely inhibited by ammonium chloride (known to neutralize intra-granular pH), whereas truncation was only partially inhibited by approximately 50%. In conclusion, a novel beta cell secretory product, des-(27-31)C-peptide, has been identified and should be considered as a potential bioactive peptide. Both truncation and degradation of C-peptide are responsible for non-equimolar secretion of insulin and C-peptide in rat beta cells.
Department of Medicine, Veterans Affairs Medical Center and University of Washington, Seattle, Washington 98108, USA. verchere@u.washington.edu
C B
CB
Verchere
M
M
Paoletta
M
M
Neerman-Arbez
K
K
Rose
J C
JC
Irminger
R L
RL
Gingerich
S E
SE
Kahn
P A
PA
Halban
D000375:Aging / Q000502:physiology*; D000595:Amino Acid Sequence; D000818:Animals; D000831:Animals, Newborn; D002096:C-Peptide / Q000096:biosynthesis* / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D002478:Cells, Cultured; D002851:Chromatography, High Pressure Liquid; D003594:Cytoplasmic Granules / Q000378:metabolism*; D006801:Humans; D007328:Insulin / Q000096:biosynthesis* / Q000737:chemistry / Q000378:metabolism; D000078790:Insulin Secretion; D007515:Islets of Langerhans / Q000254:growth & development / Q000378:metabolism*; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000096:biosynthesis* / Q000737:chemistry / Q000235:genetics; D011384:Proinsulin / Q000096:biosynthesis / Q000378:metabolism*; D011499:Protein Processing, Post-Translational; D051381:Rats; D017207:Rats, Sprague-Dawley; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry; D017386:Sequence Homology, Amino Acid; D014162:Transfection
D002096:C-Peptide; C102310:C-peptide, des-(27-31)-; D007328:Insulin; D010446:Peptide Fragments; D011994:Recombinant Proteins; D011384:Proinsulin