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X‐ray and spectrophotometric studies of the binding of proflavin to the S1 specificity pocket of human α‐thrombin
Author(s) -
Conti Elena,
Rivetti Claudio,
Wonacott Alan,
Brick Peter
Publication year - 1998
Publication title -
febs letters
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.593
H-Index - 257
eISSN - 1873-3468
pISSN - 0014-5793
DOI - 10.1016/s0014-5793(98)00235-x
Subject(s) - thrombin , protonation , chemistry , hydrogen bond , molecule , carboxylate , stereochemistry , crystallography , enzyme , biochemistry , biology , organic chemistry , ion , platelet , immunology
Proflavin can be used to study the interactions of inhibitors and substrates with thrombin by monitoring the changes in the visible absorption spectrum that occur on dye displacement. We have used microspectrophotometric methods to investigate the binding of proflavin to crystals of an α‐thrombin‐hirugen complex and have determined the structure by X‐ray crystallography. The proflavin molecule binds in the S1 pocket of the enzyme with one of the amino groups hydrogen bonded to the carboxylate of Asp‐189 while the protonated ring nitrogen is hydrogen bonded to the carbonyl of Gly‐219. This result indicates that the proflavin displacement assay can be used to specifically monitor the binding of inhibitors to the S1 pocket.