抄録
A metalloendopeptidase (MEP) isolated from rabbit liver microsomes with substrate specificity for peptides containing Arg at the P1 and P4 positions has recently proved to be identical to soluble angiotensin-binding protein present in the cytosol. Here we describe the peptide-degrading specificity of MEP, determined using various bioactive peptides and novel fluorogenic substrates for the enzyme. MEP degraded oligopeptides, including bradykinin, α-neoendorphin, bovine adrenal medulla dodecapeptide, substance P, bom-besin, neurotensin, and α-endorphin, but not polypeptides such as reduced lysozyme and histone H4, hence, MEP probably belongs to the family of endo-oligopeptidases. It cleaved most preferentially at the -Phe-Ser- bond of bradykinin(kcat/Km=2.8×104M-1 s-1) but did not cleave high molecular weight and low molecular weight kininogens, the precursors of bradykinin. MEP did not cleave angiotensin I, dynorphin A 1-13, somatostatin, and luteinizing hormone-releasing hormone, some of which are good substrates for metalloendopeptidase-24.15, metalloendopeptidase-24.16, N-arginine dibbasic convertase, and yeast endopeptidase-24.15,related peptidase. An active site-directed inhibitor of metalloendo-peptidase- 24.15, N-[1-(R,S)-carboxyl-3-phenylpropyl]-Ala-Ala-Phe-p-aminobenzoate also had no effects on the amidolytic activity of MEP. Based on the cleavage sites of bioactive peptides and processing sites of vitamin K-dependent proproteins, intramolecularly quenched fluorogenic peptide substrates were newly synthesized. Among the thirteen substrates used, the most reactive was 2-aminobenzoyl-Ala-Arg-Val-Arg-Arg-Ala-Asn-Ser-2, 4-dinitroanilinoethylamide(kcat/=9.3×105M-1S-1). An angiotensin antagonist, [Sar1, Ala8] -angiotensin II, inhibited hydrolysis of the substrate by MEP in a competitive manner (K1 = 7.6 μM).MEP cleaved oligopeptides even on the carboxyl side of proline residue and these peptides are resistant to hydrolysis by the cytosol-derived proteasome, therefore MEP may participate in the catabolism of oligopeptides in the cytosol, together with other endo-oligopeptidases.
| 元の言語 | 英語 |
|---|---|
| ページ（範囲） | 855-861 |
| ページ数 | 7 |
| ジャーナル | Journal of biochemistry |
| 巻 | 118 |
| 発行部数 | 4 |
| DOI | |
| 出版物ステータス | 出版済み - 1 1 1995 |
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All Science Journal Classification (ASJC) codes
- Biochemistry
- Molecular Biology
これを引用
Substrate specificity of rabbit liver metalloendopeptidase and its new fluorogenic peptide substrates. / Kojima, Naoko; Kawabata, Shun-Ichiro; Makinose, Yuichi; Nishino, Norikazu; Iwanaga, Sadaaki.
：: Journal of biochemistry, 巻 118, 番号 4, 01.01.1995, p. 855-861.研究成果: ジャーナルへの寄稿 › 記事
}
TY - JOUR
T1 - Substrate specificity of rabbit liver metalloendopeptidase and its new fluorogenic peptide substrates
AU - Kojima, Naoko
AU - Kawabata, Shun-Ichiro
AU - Makinose, Yuichi
AU - Nishino, Norikazu
AU - Iwanaga, Sadaaki
PY - 1995/1/1
Y1 - 1995/1/1
N2 - A metalloendopeptidase (MEP) isolated from rabbit liver microsomes with substrate specificity for peptides containing Arg at the P1 and P4 positions has recently proved to be identical to soluble angiotensin-binding protein present in the cytosol. Here we describe the peptide-degrading specificity of MEP, determined using various bioactive peptides and novel fluorogenic substrates for the enzyme. MEP degraded oligopeptides, including bradykinin, α-neoendorphin, bovine adrenal medulla dodecapeptide, substance P, bom-besin, neurotensin, and α-endorphin, but not polypeptides such as reduced lysozyme and histone H4, hence, MEP probably belongs to the family of endo-oligopeptidases. It cleaved most preferentially at the -Phe-Ser- bond of bradykinin(kcat/Km=2.8×104M-1 s-1) but did not cleave high molecular weight and low molecular weight kininogens, the precursors of bradykinin. MEP did not cleave angiotensin I, dynorphin A 1-13, somatostatin, and luteinizing hormone-releasing hormone, some of which are good substrates for metalloendopeptidase-24.15, metalloendopeptidase-24.16, N-arginine dibbasic convertase, and yeast endopeptidase-24.15,related peptidase. An active site-directed inhibitor of metalloendo-peptidase- 24.15, N-[1-(R,S)-carboxyl-3-phenylpropyl]-Ala-Ala-Phe-p-aminobenzoate also had no effects on the amidolytic activity of MEP. Based on the cleavage sites of bioactive peptides and processing sites of vitamin K-dependent proproteins, intramolecularly quenched fluorogenic peptide substrates were newly synthesized. Among the thirteen substrates used, the most reactive was 2-aminobenzoyl-Ala-Arg-Val-Arg-Arg-Ala-Asn-Ser-2, 4-dinitroanilinoethylamide(kcat/=9.3×105M-1S-1). An angiotensin antagonist, [Sar1, Ala8] -angiotensin II, inhibited hydrolysis of the substrate by MEP in a competitive manner (K1 = 7.6 μM).MEP cleaved oligopeptides even on the carboxyl side of proline residue and these peptides are resistant to hydrolysis by the cytosol-derived proteasome, therefore MEP may participate in the catabolism of oligopeptides in the cytosol, together with other endo-oligopeptidases.
AB - A metalloendopeptidase (MEP) isolated from rabbit liver microsomes with substrate specificity for peptides containing Arg at the P1 and P4 positions has recently proved to be identical to soluble angiotensin-binding protein present in the cytosol. Here we describe the peptide-degrading specificity of MEP, determined using various bioactive peptides and novel fluorogenic substrates for the enzyme. MEP degraded oligopeptides, including bradykinin, α-neoendorphin, bovine adrenal medulla dodecapeptide, substance P, bom-besin, neurotensin, and α-endorphin, but not polypeptides such as reduced lysozyme and histone H4, hence, MEP probably belongs to the family of endo-oligopeptidases. It cleaved most preferentially at the -Phe-Ser- bond of bradykinin(kcat/Km=2.8×104M-1 s-1) but did not cleave high molecular weight and low molecular weight kininogens, the precursors of bradykinin. MEP did not cleave angiotensin I, dynorphin A 1-13, somatostatin, and luteinizing hormone-releasing hormone, some of which are good substrates for metalloendopeptidase-24.15, metalloendopeptidase-24.16, N-arginine dibbasic convertase, and yeast endopeptidase-24.15,related peptidase. An active site-directed inhibitor of metalloendo-peptidase- 24.15, N-[1-(R,S)-carboxyl-3-phenylpropyl]-Ala-Ala-Phe-p-aminobenzoate also had no effects on the amidolytic activity of MEP. Based on the cleavage sites of bioactive peptides and processing sites of vitamin K-dependent proproteins, intramolecularly quenched fluorogenic peptide substrates were newly synthesized. Among the thirteen substrates used, the most reactive was 2-aminobenzoyl-Ala-Arg-Val-Arg-Arg-Ala-Asn-Ser-2, 4-dinitroanilinoethylamide(kcat/=9.3×105M-1S-1). An angiotensin antagonist, [Sar1, Ala8] -angiotensin II, inhibited hydrolysis of the substrate by MEP in a competitive manner (K1 = 7.6 μM).MEP cleaved oligopeptides even on the carboxyl side of proline residue and these peptides are resistant to hydrolysis by the cytosol-derived proteasome, therefore MEP may participate in the catabolism of oligopeptides in the cytosol, together with other endo-oligopeptidases.
UR - http://www.scopus.com/inward/record.url?scp=0028783379&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=0028783379&partnerID=8YFLogxK
U2 - 10.1093/oxfordjournals.jbchem.a124991
DO - 10.1093/oxfordjournals.jbchem.a124991
M3 - Article
C2 - 8576104
AN - SCOPUS:0028783379
VL - 118
SP - 855
EP - 861
JO - Journal of Biochemistry
JF - Journal of Biochemistry
SN - 0021-924X
IS - 4
ER -