Labeled proteinase inhibitors: versatile tools for the characterization of serine proteinases in solid-phase assays.
Peptidyl chloromethyl ketones were used for the specific labeling of proteinases by attaching a biotin group to the N-terminal end of the peptide. Such labeled peptide inhibitors allowed the detection and quantitation of proteolytic enzymes immobilized on the plastic surface of a microtiter plate, as well as on nitrocellulose. The validity of these solid-phase assays was demonstrated using subtilisin Carlsberg as a model enzyme and biotinyl-epsilon-aminocaproyl-L-alanyl-L-alanyl-L-propyl-L-phenylal++  + anyl- chloromethyl ketone as a specific reagent. In addition to being usable for the screening of a particular proteinase in a large number of samples, these assays can be adapted for the analysis of specific proteolytic enzyme present in complex mixtures.
Laboratoire de Microbiologie et Technologie Céréalières, INRA, Nantes, France.
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C
Breton-Maintier
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R
Mayer
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D
Richard-Molard
D000595:Amino Acid Sequence; D007700:Kinetics; D008969:Molecular Sequence Data; D009842:Oligopeptides / Q000494:pharmacology*; D011480:Protease Inhibitors / Q000494:pharmacology*; D012697:Serine Endopeptidases / Q000032:analysis* / Q000378:metabolism; D013381:Subtilisins / Q000032:analysis / Q000037:antagonists & inhibitors
D009842:Oligopeptides; D011480:Protease Inhibitors; D012697:Serine Endopeptidases; D013381:Subtilisins