Interaction of alpha-chymotrypsin with the fluorescent probe 1-anilinonaphthalene-8-sulfonate in solution
| Autor: | Weber Ld, El-Bayoumi Ma, Johnson Jd, Tulinsky A |
|---|---|
| Rok vydání: | 1979 |
| Předmět: |
biology
Chemistry Protein Conformation Active site Photochemistry Biochemistry Binding constant Fluorescence Anilino Naphthalenesulfonates chemistry.chemical_compound A-site Kinetics Sulfonate Spectrometry Fluorescence biology.protein Molecule Chymotrypsin Binding site Conformational isomerism Allosteric Site Protein Binding |
| Zdroj: | Biochemistry. 18(7) |
| ISSN: | 0006-2960 |
| Popis: | The binding of the fluorescence probe 1-anilinonaphthalene-8-sulfonate (Ans) to alpha-chymotrypsin (alpha-CHT) at pH 3.6 is accompanied by a dramatic enhancement of Ans fluorescence and a shift of the emission maximum to shorter wavelengths. Our study reveals that one Ans molecule binds to alpha-CHT at a site different from either the active site of alpha-CHT or the 2-p-toluidinylnapthalene-6-sulfonate binding site. the binding constant of Ans is about the same (10(4) M-1) at pH 3.6 and 6.4. Nanosecond fluorescence depolarization data indicate that Ans is rigidly bound to alpha-CHT. The fluorescence enhancement due to binding of Ans to alpha-CHT at low pH could be due to binding either to a hydrophobic site or to a site where local dipoles do not relax during the excited-state lifetime of Ans. As the pH is increased, fluorescence intensity of the Ans-alpha-CHT complex decreases appreciably; and the emission maximum shifts to longer wavelengths. The fluorescence decay curves exhibit a corresponding sensitivity to pH. The pH effect on the fluorescence of Ans-alpha-CHT can be interpreted in terms of a pH-dependent equilibrium between alpha-CHT conformers differing in the degree of mobility of polar residues and water molecules at the Ans binding site or structural changes in the Ans binding site. |
| Databáze: | OpenAIRE |
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