Abstract
A new chromogenic substrate and two inhibitors with the common peptide sequence X-Ala-Ala-Phe-Y have been synthesized, and were found to be of higher efficiency as hitherto available customary substrates and inhibitors for thermitase, subtilisin BPN' and alpha-chymotrypsin. The proteolytic coefficient kcat/Km for the hydrolysis of the substrate Suc-Ala-Ala-Phe-pNA is about 80 times higher in the case of thermitase and subtilisin BPN' and about 20 times higher for alpha-chymotrypsin compared with Suc-Ala3-pNA and Suc-Phe-pNA, respectively. The irreversible inhibitory effect of Z-Ala2-Phe-CH2Cl compared with Z-Phe-CH2Cl is 280 times greater for thermitase and subtilisin BPN' and 50 times for alpha-chymotrypsin. The corresponding methyl ketone Z-Ala2-Phe-CH3 is a high affinity competitive inhibitor for thermitase but not for the two other enzymes.
MeSH terms
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Amino Acid Chloromethyl Ketones / metabolism
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Amino Acid Chloromethyl Ketones / pharmacology
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Amino Acid Sequence
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Binding Sites
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Binding, Competitive
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Chromogenic Compounds / metabolism
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Chromogenic Compounds / pharmacology
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Chymotrypsin / antagonists & inhibitors
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Chymotrypsin / metabolism*
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Endopeptidases / metabolism*
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In Vitro Techniques
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Kinetics
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Oligopeptides / metabolism
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Oligopeptides / pharmacology
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Protease Inhibitors
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Serine Endopeptidases*
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Substrate Specificity
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Subtilisins / antagonists & inhibitors
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Subtilisins / metabolism*
Substances
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Amino Acid Chloromethyl Ketones
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Chromogenic Compounds
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Oligopeptides
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Protease Inhibitors
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benzyloxycarbonylalanyl-alanyl-phenylalanine-4-nitroanilide
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succinyl-alanyl-alanyl-phenylalanine-4-nitroanilide
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benzyloxycarbonylalanyl-alanyl phenylalanine chloromethyl ketone
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Endopeptidases
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Serine Endopeptidases
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Subtilisins
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thermitase
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Chymotrypsin