Interaction of 7-hydroxy-8-(phenylazo)1,3-naphthalenedisulfonate with bovine plasma albumin. Spectroscopic studies.
Interaction of Orange G (OG) with bovine plasma albumin (BPA) has been investigated using NMR, UV-visible absorption, CD, and fluorescence techniques. The bound conformation of OG is a compact structure with N9-N10 bond in a non-planar syn conformation. The binding causes a decrease in the 478-nm absorption band of OG. The analysis of the binding isotherm generated from UV-visible absorption measurements gives a dissociation constant of 10 microM and stoichiometry 1:1 for BPA.OG complex. Dissociation constant is invariant in the pH range 5.0-8.0 and is approximately 20 times higher at pH 4.0 than its value at pH 7.0. Near and far UV-CD studies indicate alterations in the helical content and in the tertiary structure of the protein on complexation. The binding induces (-) and (+) CD at 335 nm and 465 nm, respectively. The binding also results into an increase in the steady state fluorescence anisotropy of OG without affecting emission maximum and quantum yield. Fluorescence data indicate that quenching of Trp fluorescence by OG is static in nature and OG selectively binds near Trp-135. Observation of similar rotational correlation time for BPA and BPA.OG complex indicates that the overall globular structure of BPA remains unaltered on binding despite certain internal rearrangement in the protein structure.
Department of Chemical Sciences, Tata Institute of Fundamental Research, Homi Bhabha Road, Navy Nagar, Colaba, Mumbai 400 005, India.
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Patel
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Srivastava
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Phadke
D000818:Animals; D001391:Azo Compounds / Q000737:chemistry* / Q000493:pharmacokinetics*; D001665:Binding Sites; D002417:Cattle; D002942:Circular Dichroism; D007700:Kinetics; D009682:Magnetic Resonance Spectroscopy; D008968:Molecular Conformation; D011487:Protein Conformation; D012710:Serum Albumin, Bovine / Q000737:chemistry* / Q000378:metabolism*; D013050:Spectrometry, Fluorescence; D013053:Spectrophotometry
D001391:Azo Compounds; C008710:Orange G; D012710:Serum Albumin, Bovine