Hydrolysis by somatic angiotensin-I converting enzyme of basic dipeptides from a cholecystokinin gastrin and a LH-RH peptide extended at the C-terminus with Gly-Arg/Lys-Arg, but not from diarginyl insulin

Endoproteolytic cleavage of protein prohormones often generates intermediat es extended at the C-terminus by Arg-Arg or Lys-Arg, the removal of which b y a carboxypeptidase (CPE) is normally an important step in the maturation of many peptide hormones. Recent studies in mice that lack CP activity indi cate the existence of alternative tissue or plasma enzymes capable of remov ing C-terminal basic residues from prohormone intermediates. Using inhibito rs of angiotensin I-converting enzyme (ACE) and CP, we show that both these enzymes in mouse serum can remove the basic amino acids from the C-terminu s of CCK5-GRR and LH-RH-GKR, but only CP is responsible for converting diar ginyl insulin to insulin. ACE activity removes C-terminal dipeptides to gen erate the Gly-extended peptides, whereas CP hydrolysis gives rise to CCK5-G R and LH-RH-GK, both of which are susceptible to the dipeptidyl carboxypept idase activity of ACE. Somatic ACE has two similar protein domains (the N-d omain and the C-domain), each with an active site that can display differen t substrate specificities. CCK5-GRR is a high-affinity substrate for both t he N-domain and C-domain active sites of human sACE (K-m of 9.4 mu M and 9. 0 mu M, respectively) with the N-domain showing greater efficiency (k(cat): K-m ratio of 2.6 in favour of the N-domain). We conclude that somatic forms of ACE should be considered as alternatives to CPs for the removal of basi c residues from some Arg/Lys-extended peptides.