Proglucagon processing in porcine and human pancreas.

Abstract

In the pancreas proglucagon (PG), a peptide precursor of 160 amino acids is cleaved to produce glucagon and a 30-amino acid N-terminal flanking peptide, but the fate of the C-terminal flanking peptide (99 amino acids) is incompletely known.We subjected acid ethanol extracts of human and porcine pancreases to gel filtration and analyzed the fractions with specific radioimmunoassays for the following regions of proglucagon: PG 62-69, PG 72-81, PG 78-87, PG 98-107 amide, PG 126-134, and PG 142)-158.Based on these assays and successive purifications by high performance liquid chromatography we isolated and purified to homogeneity three porcine peptides which were subjected to mass spectrometry and sequencing.One peptide was PG 64-69.The second was PG 72-108, as determined by mass spectrometry, N-terminal amino acid sequencing, and specific radioimmunoassays.The third had a molecular size of approximately lO, OOO, an N-terminal sequence corresponding to PG 72-81, and a C-terminal sequence terminating at PG 158 (specific radioimmunoassay).A similar peptide with an identical N-terminal sequence, a C-terminal sequence corresponding to PG 146-158, and a molecular mass of 9969 (theoretical mass for human PC 72-158 = 9971) was isolated from human pancreas together with small amounts of a peptide corresponding to PG 72-107 amide.Thus, the pancreatic processing of the C-terminal flanking peptide in proglucagon includes the formation of equimolar (to glucagon) amounts of PG 64-69 and PG 72-158 (major proglucagon fragment) and smaller amounts of N-terminally extended glucagon-like peptide-1 (GLP-1) (PG 72-108 in pigs and PG 72-107 amide in humans).In the pancreas the glucoregulatory hormone, glucagon, is produced by cleavage of a 160-amino acid precursor peptide, proglucagon (PG),' the structure of which has been deduced by sequence analysis of coding polynucleotides (1-3) (Fig. 1).The N-terminal part of porcine proglucagon consists of glicentin, a peptide of 69-amino acid residues, first isolated from endocrine cells in the intestinal mucosa (4), in which the single glucagon gene is also expressed (5).In glicentin (and thus in progluca-* This study was supported by grants from the Danish Medical Research Council, the Biotechnology Programme for Signal Peptides, the Lundbeck Foundation, and Brdr.Hartmann's Foundation.The costs of publication of this article were defrayed in part by the payment of page in accordance with 18 U.S.C.Section 1734 solely to indicate this fact.