Characterisation of endothelin converting enzyme-1 shedding from endothelial cells.
The aim of this study was to determine if endothelin converting enzyme-1 (ECE-1) like other members of this metalloprotease family undergoes ectodomain shedding. The release/shedding of catalytically active ECE-1 was measured by monitoring the fluorescence resulting from the cleavage of a specific quenched fluorescent substrate. Catalytically active ECE-1 was detected in the media of human umbilical vein endothelial cells, and was confirmed by mass spectrometry based assays. Specificity of cleavage was confirmed by using both narrow and broad specificity inhibitors. In conclusion we demonstrate and characterize for the first time, ECE-1 shedding from the surface of endothelial cells.
Kuruppu
Sanjaya
S
Department of Biochemistry and Molecular Biology, Monash University, Vic 3800, Australia. Sanjaya.Kuruppu@med.monash.edu.au
Reeve
Shane
S
Ian Smith
A
A
D000595:Amino Acid Sequence; D016282:Aspartic Acid Endopeptidases / Q000737:chemistry / Q000378:metabolism*; D002384:Catalysis / Q000187:drug effects; D002460:Cell Line; D002853:Chromatography, Liquid; D004151:Dipeptides / Q000494:pharmacology; D042783:Endothelial Cells / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D000072396:Endothelin-Converting Enzymes; D016232:Endothelins / Q000378:metabolism; D004789:Enzyme Activation / Q000187:drug effects; D006020:Glycopeptides / Q000494:pharmacology; D006801:Humans; D008666:Metalloendopeptidases / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000378:metabolism*; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D013379:Substrate Specificity; D053719:Tandem Mass Spectrometry; D015244:Thiorphan / Q000494:pharmacology; D014471:Umbilical Veins / Q000166:cytology
D004151:Dipeptides; D016232:Endothelins; D006020:Glycopeptides; C078131:N-(2(R)-2-(hydroxamidocarbonylmethyl)-4-methylpentanoyl)-L-tryptophan methylamide; D015244:Thiorphan; D016282:Aspartic Acid Endopeptidases; D008666:Metalloendopeptidases; C000606923:ECE1 protein, human; D000072396:Endothelin-Converting Enzymes; C008890:phosphoramidon