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  • 标题:Two Molecular Species of Oxytocinase (L-Cystine Aminopeptidase) in Human Placenta : Purification and Characterization
  • 本地全文:下载
  • 作者:Chie ITOH ; Maho WATANABE ; Atsuo NAGAMATSU
  • 期刊名称:Biological and Pharmaceutical Bulletin
  • 印刷版ISSN:0918-6158
  • 电子版ISSN:1347-5215
  • 出版年度:1997
  • 卷号:20
  • 期号:1
  • 页码:20-24
  • DOI:10.1248/bpb.20.20
  • 出版社:The Pharmaceutical Society of Japan
  • 摘要:Two different forms of oxytocinase (L-cystine aminopeptidase, CAP; EC 3.4.11.3) were purified from the 9000 g and 105000 g precipitate fractions of human placenta homogenate by sequential chromatography on columns of hydroxyapatite, DE-32, nickel ion affinity, and Sephadex G-200. One species (CAP-I) purified from the mitochondrial/lysosomal fraction migrated on sodium dodecyl sulfate (SDS)-polyacrylamide gel electrophoresis with an apparent molecular mass of 61 kDa; the other (CAP-II) from the microsomal fraction was composed of two subunits with molecular masses of 56 and 40 kDa. The molecular masses of CAP-I and CAP-II estimated by gel filtration were 64 and 97 kDa, respectively. The specific activities of the two species for S-benzyl-L-cysteine p-nitroanilide increased by 357- (for CAP-I) and 139-fold (for CAP-II) compared with the starting preparations. The optimal pH values toward the artificial substrate were approx. 7.4-8.0 for CAP-I and 6.8-8.0 for CAP-II. The Km and Vmax values toward oxytocin were 5.6 μM and 23.4 μmol/h/mg protein for CAP-I, and 38μM and 15.6 μmol/h/mg protein for CAP-II. Both enzymes were inhibited by the metal-chelating agents, EDTA and o-phenanthroline, whereas they were specifically activated by addition of Co2+ : CAP-I was more sensitive to these reagents than CAP-II. L-Methionine strongly inhibited CAP-I, while CAP-II activity was only slightly affected. CAP-II was more sensitive to amastatin than CAP-I. Thus, the two enzymes are quite distinct in their molecular nature and biochemical properties. They may play a regulatory role in the metabolism of oxytocin and other biologically active peptides in intact placenta.
  • 关键词:cystinyl aminopeptidase;oxytocin;vasopressin;purification;human placenta
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