Isolation of products and intermediates of pancreatic prosomatostatin processing: use of fast atom bombardment mass spectrometry as an aid in analysis of prohormone processing

Publisher Website
Google Scholar
Abstract
Major products and an intermediate in the proteolytic processing pathway of preprosomatostatin I from anglerfish (Lophius americanus) were purified and characterized. Proteolytic mapping by fast atom bombardment mass spectrometry was used to rapidly locate regions of the peptides whose masses deviated from those deduced from the cDNA sequence. Amino acid analysis and partial Edman sequencing were also used to confirm the structures. The protein structural data indicate a Glu for Gly substitution at position 83 of preprosomatostatin I (aPPSS-I, numbering from the initiator Met) relative to the cDNA sequence. Two of the peptides isolated, aPPSS-I (26-52) (7.5 nmol .cntdot. g-1) and aPPSS-I (26-92) (49.5 nmol .cntdot. g-1), define signal cleavage as occurring between Cys-25 and Ser-26. A partial sequence was obtained from fragment ions in the mass spectrum of a peptide corresponding to aPPSS-I (94-105) (58 nmol .cntdot. g-1). The 14-residue somatostatin [Ss-14 corresponding to aPPSS-I (108-121)] has previously been isolated [Noe, B. D., Spiess, J., Rivier, J. E., and Vale, W. (1979) Endocrinology (Baltimore) 105, 1410-1415]. Taken together, these peptides suggest a pathway for prosomatostatin I processing in which the residues corresponding to SS-14 and the immediately preceding 14 residues are cleaved from the prohormone via endoproteolysis (order of cleavage not determined). The fragment aPPSS-I (94-105) was isolated in lower yield than SS-14 and may represent a secondary site of cleavage. Subsequent cleavage at arginine-53 results in the minor peptide aPPSS-I (26-52). The terminal basic amino acids generated by endoproteolytic processing were not found for any of the peptides isolated. The peptides described were identified as products of aPPSS-I processing in radiolabeling studies using intact anglerfish islets [Noe, B. D., Andrews, P. C., Dixon, J. E., and Spiess, J. (1986) J. Cell Biol. 103, 1205-1211].