Characterization of endothelin converting enzyme activities in soluble fraction of bovine cultured endothelial cells.
Endothelin converting enzyme activities in the soluble fraction of cultured bovine aortic endothelial cells were characterized. The two major endothelin converting enzyme activities were eluted from a hydrophobic chromatography column and the elution profile of the endothelin converting enzyme activities was the same as that of cathepsin D activities. These activities had a same pH optimum at pH 3.5 and were effectively inhibited by pepstatin A. Furthermore, anti-cathepsin D antiserum absorbed these activities as well as cathepsin D activity. Immunoblotting analysis using the antiserum showed the major active fractions have immunostainable components of identical molecular weights with cathepsin D. From these results, we concluded that the major endothelin converting activities in the soluble fraction of endothelial cells are due to cathepsin D. In addition to these cathepsin D activities, a minor endothelin converting enzyme activity with an optimum pH at 3.5 was found, which does not have angiotensin I generating (cathepsin D) activity from renin substrate and needs much higher concentrations of pepstatin A to inhibit the activity than cathepsin D.
Department of Biochemistry, University of Tsukuba, Ibaraki, Japan.
T
T
Sawamura
S
S
Kimura
O
O
Shinmi
Y
Y
Sugita
M
M
Kobayashi
Y
Y
Mitsui
M
M
Yanagisawa
K
K
Goto
T
T
Masaki
D000818:Animals; D016282:Aspartic Acid Endopeptidases; D002402:Cathepsin D / Q000037:antagonists & inhibitors / Q000378:metabolism*; D002417:Cattle; D010450:Endopeptidases / Q000378:metabolism; D016232:Endothelins; D004730:Endothelium, Vascular / Q000201:enzymology*; D006863:Hydrogen-Ion Concentration; D007158:Immunologic Techniques; D008970:Molecular Weight; D010455:Peptides / Q000378:metabolism*; D012995:Solubility
D016232:Endothelins; D010455:Peptides; D010450:Endopeptidases; D016282:Aspartic Acid Endopeptidases; D002402:Cathepsin D