Characterization of azidobenzamidines as photoaffinity labels for trypsin

Meta- and para-azidobenzamidine have been prepared and evaluated as photoaffinity labels. The compounds inhibit trypsin reversible in the dark and are competitive with substrate binding. Upon photolysis, irreversible noncompetitive inhibition is observed and is dependent upon concentration, photolys...

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Bibliographic Details
Published in:The Journal of biological chemistry 1978-03, Vol.253 (6), p.1846-1852
Main Authors: DeTraglia, M C, Brand, J S, Tometsko, A M
Format: Article
Language:English
Subjects:
Affinity Labels - chemical synthesis
Affinity Labels - pharmacology
Amidines - pharmacology
Azides - pharmacology
Benzamidines - pharmacology
Kinetics
Photolysis
Spectrophotometry, Ultraviolet
Trypsin - metabolism
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Summary:Meta- and para-azidobenzamidine have been prepared and evaluated as photoaffinity labels. The compounds inhibit trypsin reversible in the dark and are competitive with substrate binding. Upon photolysis, irreversible noncompetitive inhibition is observed and is dependent upon concentration, photolysis time, and pH. Specificity of the probes is indicated by experiments in which p-tosyl-l-arginine methyl ester, a trypsin substrate, is used to protect against photoinactivation. Maximum inactivation of trypsin is achieved at pH 6.2 using either azidobenzamidine derivative. Evaluation of the pH dependence of photoaffinity labeling may provide a sensitive tool for probing conformational changes in inhibitor binding sites. These studies provide a basis for the use of azidobenzamidines as photoaffinity analogs of lysine and arginine side chains.
ISSN:0021-9258
1083-351X
DOI:10.1016/S0021-9258(19)62329-X