title: Enzymes that process somatostatin precursors. A novel endoprotease that cleaves before the arginine-lysine doublet is involved in somatostatin-28 convertase activity of rat brain cortex.
abstract: The selective processing activity which generates both the NH2- and COOH-terminal fragments of the octacosapeptide somatostatin-28 (S-28) was investigated. Separation into two distinct proteolytic activities was achieved by ion-exchange chromatography. An endoprotease cleaving either the substrate Pro-Arg-Glu-Arg-Lys-Ala-Gly-Ala-Lys-Asn-Tyr-NH2, i.e. [Ala17,Tyr20]S-28-(10-20)-NH2 (peptide I), or the octacosapeptide somatostatin-28, on the NH2 side of the Arg-Lys doublet was separated from an aminopeptidase B-like activity. Whereas the endoprotease cleaves a single peptide bond, between Glu12 and Arg13 of S-28, the aminopeptidase B-like enzyme removes both Arg13 and Lys14 stepwise from the NH2 terminus of the corresponding COOH-terminal fragment. This endoprotease activity peaks around pH 8.5, whereas the optimal aminopeptidase B-like activity is in the pH range 6.2-8.5. Combination of both enzymes resulted in the recovery of the overall S-28 convertase activity with an optimal pH at 7. In addition, this endoprotease appears to be very sensitive to divalent cations since it is strongly inhibited by chelating agents. The use of selectively modified undecapeptides derived from the reference substrate peptide I by a single modification of the amino acids Glu12, Arg13, and Lys14 at the cleavage locus showed that both basic residues are critically important, whereas Glu12 is not. It is proposed that S-28 processing involves a divalent cation-sensitive endoprotease that is sensitive to thiol reagents, which cleaves before the Arg-Lys doublet, which is not trypsin-like, and whose action is coupled to an aminopeptidase B-like enzyme.
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001120:Arginine; D002540:Cerebral Cortex / Q000201:enzymology*; D010450:Endopeptidases / Q000378:metabolism*; D007700:Kinetics; D008239:Lysine; D051381:Rats; D013004:Somatostatin / Q000096:biosynthesis*; D013379:Substrate Specificity
chemical_list: D013004:Somatostatin; D001120:Arginine; D010450:Endopeptidases; C045514:somatostatin-28 convertase; D008239:Lysine