2108-P: Complex Posttranslational Processing of Glucagon Prohormone in Human Pancreatic Islets

An array of signaling peptides and peptide hormones in pancreatic islets results from posttranslational processing of precursor proteins by well-characterized endopeptidases. While the encoded peptide complement can be predicted in silico from the prohormone sequence, actual peptide products often are more diverse due to additional chemical or enzymatic modifications of mature predicted peptides. Using liquid chromatography-tandem mass spectrometry (LC-MS/MS) for comprehensive molecular characterization, we compiled an experimentally deduced database of endogenous peptides in human pancreatic islets and revealed an unexpected complex processing of several prohormones including proglucagon. Live pancreatic islets from several donors were provided by the Human Pancreas Analysis Program, University of Pennsylvania. Total of 2034 chemically unique peptide sequences were identified and matched to 477 proteins in the human protein database. Among interesting observations, we find differential cleavage of the signal peptide on the glucagon precursor protein, NP_002045.1, and unusual further processing of its encoded mature peptides. Alternate signal peptide cleavage sites are confirmed by detection of both classical GRPP, R[21]-D[50], and its novel variant S[18]-D[50]. An amidated IP-2 D[131]-L[142] and an isoform of MPGF D[131]-D[178], missing the GLP-1 sequence were detected. Previously unreported N-terminus acetylation of glucagon, GLP-1, GLP-1 (7-36), and oxyntomodulin was detected. Novel peptides potentially resulted from variable processing of mature glucagon, GRPP, and oxyntomodulin at internal convention cleavage sites were characterized. New chemically unique peptides originating from proglucagon and documented in this study indicate a more complex posttranslational processing of proglucagon in human islets than previously thought. The finding expands opportunity for mechanistic studies on pathology, progression, and treatments of diabetes.