Quenching of the intrinsic fluorescence of bovine serum albumin by chlorpromazine and hemin.
The binding of chlorpromazine (CPZ) and hemin to bovine serum albumin was studied by the fluorescence quenching technique. CPZ is a widely used anti-psychotic drug that interacts with blood components, influences bioavailability, and affects function of several biomolecules. Hemin is an important ferric residue of hemoglobin that binds within the hydrophobic region of albumin with high specificity. Quenching of the intrinsic fluorescence of bovine serum albumin (BSA) was observed by selectively exciting tryptophan residues at 290 nm. Emission spectra were recorded in the range from 300 to 450 nm for each quencher addition. Stern-Volmer graphs were plotted, and the quenching constant estimated for BSA solution titrated with hemin at 25 masculine C was 1.44 (+/- 0.05) x 10(5) M(-1). Results showed that bovine albumin tryptophans are not equally accessible to CPZ, in agreement with the idea that polar or charged quenchers have more affinity for amino acid residues on the outer wall of the protein. Hemin added to albumin solution at a molar ratio of 1:1 quenched about 25% of their fluorescence. The quenching effect of CPZ on albumin-hemin solution was stronger than on pure BSA. This increase can be the result of combined conformational changes in the structure of albumin caused firstly by hemin and then by CPZ. Our results suggest that the primary binding site for hemin on bovine albumin may be located asymmetrically between the two tryptophans along the sequence formed by subdomains IB and IIA, closer to tryptophan residue 212.
Silva
D
D
Departamento de Ciências Fisiológicas, Universidade do Estado do Rio Janeiro, Rio de Janeiro, RJ, Brazil. dilsons@uerj.br
Cortez
C M
CM
Louro
S R W
SR
D000818:Animals; D014150:Antipsychotic Agents / Q000494:pharmacology*; D002417:Cattle; D002746:Chlorpromazine / Q000494:pharmacology*; D004347:Drug Interactions; D006427:Hemin / Q000494:pharmacology*; D011485:Protein Binding; D012710:Serum Albumin, Bovine / Q000737:chemistry* / Q000187:drug effects; D013050:Spectrometry, Fluorescence; D014364:Tryptophan / Q000032:analysis
D014150:Antipsychotic Agents; D012710:Serum Albumin, Bovine; D006427:Hemin; D014364:Tryptophan; D002746:Chlorpromazine