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 intermediates extended at the C-terminus by Arg-Arg or Lys-Arg, the removal of which by a carboxypeptidase (CPE) is normally an important step in the maturation of many peptide hormones. Recent studies in mice that lack CP activity indicate the existence of alternative tissue or plasma enzymes capable of removing C-terminal basic residues from prohormone intermediates. Using inhibitors 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-terminus of CCK5-GRR and LH-RH-GKR, but only CP is responsible for converting diarginyl insulin to insulin. ACE activity removes C-terminal dipeptides to generate the Gly-extended peptides, whereas CP hydrolysis gives rise to CCK5-GR and LH-RH-GK, both of which are susceptible to the dipeptidyl carboxypeptidase activity of ACE. Somatic ACE has two similar protein domains (the N-domain and the C-domain), each with an active site that can display different substrate specificities. CCK5-GRR is a high-affinity substrate for both the N-domain and C-domain active sites of human sACE (Km of 9.4 microm and 9.0 microm, respectively) with the N-domain showing greater efficiency (kcat : Km 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 basic residues from some Arg/Lys-extended peptides.
Isaac
R E
RE
School of Biology, University of Leeds, UK. r.e.isaac@leeds.ac.uk
Michaud
A
A
Keen
J N
JN
Williams
T A
TA
Coates
D
D
Wetsel
W C
WC
Corvol
P
P
D000818:Animals; D001120:Arginine / Q000737:chemistry; D002766:Cholecystokinin / Q000737:chemistry*; D004151:Dipeptides / Q000378:metabolism*; D005260:Female; D005755:Gastrins / Q000737:chemistry*; D005998:Glycine / Q000737:chemistry; D007987:Gonadotropin-Releasing Hormone / Q000737:chemistry*; D006801:Humans; D006868:Hydrolysis; D007328:Insulin / Q000031:analogs & derivatives* / Q000737:chemistry; D008239:Lysine / Q000737:chemistry; D051379:Mice; D007703:Peptidyl-Dipeptidase A / Q000378:metabolism*; D013379:Substrate Specificity
D004151:Dipeptides; D005755:Gastrins; D007328:Insulin; D007987:Gonadotropin-Releasing Hormone; C063853:insulin, Arg(B31,B32)-; D002766:Cholecystokinin; D001120:Arginine; D007703:Peptidyl-Dipeptidase A; D008239:Lysine; D005998:Glycine