The Hemoregulatory Peptide N-Acetyl-Ser-Asp-Lys-Pro Is a Natural and Specific Substrate of the N-terminal Active Site of Human Angiotensin-converting Enzyme (*)
- From the Centre National de la Recherche Scientifique, Institut de Chimie des Substances Naturelles, 91198 Gif-sur-Yvette, France and the
- Institut National de la Santé et de la Recherche Médicale, Unit 36, College de France, 3 rue d'Ulm 75005 Paris, France
- ¶ To whom correspondence should be addressed: College de France, 3 rue d'Ulm, 75005 Paris France. Tel.: 33-1-44-27-16-75; Fax: 33-1-44-27-16-91.
Abstract
Angiotensin I-converting enzyme (ACE) is a zinc-dipeptidyl carboxypeptidase, which contains two similar domains, each possessing
a functional active site. Respective involvement of each active site in the degradation of the circulating peptide N-acetyl-seryl-aspartyl-lysyl-proline (AcSDKP), a negative regulator of hematopoietic stem cell proliferation, was studied
by using wild-type recombinant ACE and two full-length mutants containing a single functional site. Both the N- and C-active
sites of ACE exhibit dipeptidyl activity toward AcSDKP, with K
values of 31 and 39 μM, respectively. However, the N-active site hydrolyzes the peptide 50 times faster compared with the
C-active site, with k
/K
values of 0.5 and 0.01 μM
•s
, respectively. The predominant role of the N-active site in AcSDKP hydrolysis was confirmed by the inhibition of hydrolysis
using a monoclonal antibody specifically directed against the N-active site. The N-domain specificity for AcSDKP will aid
the identification of specific inhibitors for this domain. This is the first report of a highly specific substrate for the
N-active site of ACE, with kinetic constants in the range of physiological substrates, suggesting that ACE might be involved
via its N-terminal active site in the in vivo regulation of the local concentration of this hemoregulatory peptide.
Footnotes
-
↵* This work was supported by the Ligue Nationale contre le Cancer, by the Institut National de la Santé et de la Recherche Médicale, and by a grant from the Bristol-Myers-Squibb Institute for Medical Research. The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore by hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
-
↵1 The abbreviations used are:
- ACE
-
angiotensin-converting enzyme
- LH-RH
-
luteinizing hormone-releasing hormone
- CHO
-
Chinese hamster ovary
- Hip
-
hippuryl
- HPLC
-
high performance liquid chromatography
- mAb
-
monoclonal antibody
- AcSDKP
-
N-acetyl-seryl-aspartyl-lysyl-proline.
-
↵2A. Rousseau, A., Michaud, M.-T. Chauvet, M. Lenfant, and P. Corvol, unpublished results.
-
↵3D. Guillaume, unpublished results.
-
- Received August 7, 1994.
- Revision received November 7, 1994.
- © 1995 by The American Society for Biochemistry and Molecular Biology, Inc.
