title: Interaction of terbium and calcium with chicken cystatin.
abstract: The emission intensity of the fluorescent lanthanide, terbium, is shown to be enhanced upon binding to chicken cystatin. Fluorescence titrations indicate the presence of a single high affinity binding site per molecule. Binding of the terbium results in a 29% quenching of the fluorescence of the single tryptophan residue in the molecule. Calcium displaces the terbium from cystatin as judged by the decrease of terbium fluorescence in competition titrations. Similar titrations with magnesium or strontium demonstrate that the metal binding site of cystatin exhibits specificity for calcium or terbium. Analysis of the N-terminal sequence of chicken cystatin suggests the presence of a putative consensus sequence for a metal binding site between residues 13 and 24. Calcium causes a 17% decrease in the tryptophan fluorescence of cystatin, indicating that an induced conformational change accompanies metal binding. The increased quenching observed with terbium appears to be the result of resonance energy transfer from tryptophan to terbium. From the critical distance for energy transfer from tryptophan to terbium, it is estimated that the terbium binding site lies approximately 12 A from the single tryptophan residue in the molecule.
authors.affiliation: Department of Biochemistry, University of Rochester Medical Center, New York 14642.
authors.forename: E T
authors.initials: ET
authors.lastname: Bell
authors.forename: J D
authors.initials: JD
authors.lastname: Featherstone
authors.forename: J E
authors.initials: JE
authors.lastname: Bell
mesh_terms: D000818:Animals; D001665:Binding Sites; D002118:Calcium / Q000378:metabolism*; D002645:Chickens; D015891:Cystatins; D004735:Energy Transfer; D006801:Humans; D011487:Protein Conformation; D011506:Proteins / Q000378:metabolism*; D055333:Salivary Cystatins; D012471:Salivary Proteins and Peptides; D013050:Spectrometry, Fluorescence; D013725:Terbium / Q000378:metabolism*; D014364:Tryptophan / Q000378:metabolism
chemical_list: D015891:Cystatins; D011506:Proteins; D055333:Salivary Cystatins; D012471:Salivary Proteins and Peptides; C100158:cystatin, egg-white; D013725:Terbium; D014364:Tryptophan; D002118:Calcium