ICCAS OpenIR
Characterization of local polarity and structure of Cys121 domain in beta-lactoglobulin with a new thiol-specific fluorescent probe
Wang, Xiaochun; Wang, Shujuan; Ma, Huimin
2008
Source PublicationANALYST
ISSN0003-2654
Volume133Issue:4Pages:478-484
AbstractThe design and synthesis of a new polarity-sensitive fluorescent probe, 3-(4-chloro-6-p-maleimidylphenoxyl-1,3,5- triazinylamino)-7-dimethylamino-2-methylphenazine, is reported for characterizing the local polarity and structure, such as the thiol domain, of a protein. The probe comprises a polarity-sensitive fluorophore (neutral red moiety) and a thiol-specific labeling group (maleimidyl moiety). The probe exhibits a sensitive response of shift of fluorescence maximum emission wavelength to solvent polarity, but not to pH and temperature, which makes the probe suitable for determining the local polarity change of a protein denatured by pH or temperature. The application of this kind has been first demonstrated for the polarity detection of the Cys121 domain of beta-lactoglobulin. It is found that the polarity of the Cys121 domain corresponds to a dielectric constant of 17.3, and this value hardly alters after heat treatment, which may be attributed to the improved thermal reversibility by the Cys121 modi. cation. The simple mixture of the probe and the protein at pH 5.6 is also studied, revealing that the free probe prefers to bind to an outer hydrophobic region. Heat treatment of the mixture causes the modi. cation of Cys121 residues; this modi. cation does not completely destroy the calyx but results in the opening of a channel for the probe to enter the calyx of b-lactoglobulin. These results show that Cys121 plays an important role not only in the thermal reversibility but also in the accessibility of the calyx to a ligand. The strategy presented here further indicates that the combination of polarity-sensitive fluorescence probe with site-specific labeling may serve as a powerful means for elucidating structures and properties of proteins.
DOI10.1039/b717230c
Indexed BySCI
Language英语
WOS IDWOS:000254315600010
PublisherROYAL SOC CHEMISTRY
Citation statistics
Cited Times:12[WOS]   [WOS Record]     [Related Records in WOS]
Document Type期刊论文
Identifierhttp://ir.iccas.ac.cn/handle/121111/63642
Collection中国科学院化学研究所
Corresponding AuthorMa, Huimin
AffiliationChinese Acad Sci, Beijing Natl Lab Mol Sci, Inst Chem, Beijing 100080, Peoples R China
Recommended Citation
GB/T 7714
Wang, Xiaochun,Wang, Shujuan,Ma, Huimin. Characterization of local polarity and structure of Cys121 domain in beta-lactoglobulin with a new thiol-specific fluorescent probe[J]. ANALYST,2008,133(4):478-484.
APA Wang, Xiaochun,Wang, Shujuan,&Ma, Huimin.(2008).Characterization of local polarity and structure of Cys121 domain in beta-lactoglobulin with a new thiol-specific fluorescent probe.ANALYST,133(4),478-484.
MLA Wang, Xiaochun,et al."Characterization of local polarity and structure of Cys121 domain in beta-lactoglobulin with a new thiol-specific fluorescent probe".ANALYST 133.4(2008):478-484.
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