title: Site-selective probe for investigating the asynchronous unfolding of domains in bovine serum albumin.
abstract: A convenient method is proposed for precise investigation of the asynchronous structural transition of the domains in bovine serum albumin (BSA) during unfolding process. The method is based on a site-selective probe, alizarin red S (ARS), which has a high affinity to the subdomain IIA of BSA. BSA-ARS complex was formed and gradually unfolded by urea from 0 to 8.0M. The unfolding occurred in different domains of BSA resulted in distinct alterations of the microenvironment of the bound ARS. The spectral response of BSA-ARS complex, including the color, the UV absorption at 530 and 432 nm, and the intrinsic fluorescence at 342 and 310 nm with the excitation wavelength of 280 nm, showed slight changes in the urea concentration from 0 to 4.5M, drastic changes from 4.5 to 6.0M, and almost no changes from 6.0 to 8.0M. The redox behavior of bound ARS between 0.3 and 0.8 V also showed the same trend. Consequently, a two-step, three-state transition process was monitored by naked eyes, UV-vis spectroscopy and electrochemistry. It is the first report to realize the indicator of the intermediate state during the unfolding process of BSA through convenient methods instead of expensive approaches. The work provides a facile method for the investigation of the unfolding process of multidomain proteins.
authors.affiliation: School of Chemistry and Chemical Engineering, Sun Yat-Sen University, Guangzhou 510275, PR China. wuhai317@126.com
authors.forename: Hai
authors.initials: H
authors.lastname: Wu
authors.forename: Po
authors.initials: P
authors.lastname: Wang
authors.forename: Xiao
authors.initials: X
authors.lastname: Hu
authors.forename: Zong
authors.initials: Z
authors.lastname: Dai
authors.forename: Xiaoyong
authors.initials: X
authors.lastname: Zou
mesh_terms: D015335:Molecular Probes; D011489:Protein Denaturation; D012710:Serum Albumin, Bovine / Q000737:chemistry*; D013050:Spectrometry, Fluorescence; D013056:Spectrophotometry, Ultraviolet
chemical_list: D015335:Molecular Probes; D012710:Serum Albumin, Bovine