期刊
ARCHIVES OF BIOCHEMISTRY AND BIOPHYSICS
卷 468, 期 1, 页码 15-21出版社
ELSEVIER SCIENCE INC
DOI: 10.1016/j.abb.2007.09.011
关键词
tear lipocalin; time-resolved fluorescence; binding sites; ANS; heterogeneous binding; lipocalin-1; von Ebner's gland protein; human tear protein
资金
- NEI NIH HHS [EY 00331, R29 EY011224, P30 EY000331, EY 11224, R01 EY011224-12, R01 EY011224] Funding Source: Medline
8-Anilino-1-naphthalenesulfonic acid (ANS) is widely used as a probe for locating binding sites of proteins. To characterize the binding sites of tear lipocalin (TL), we studied ANS binding to apoTL by steady-state and time-resolved fluorescence. Deconvolution of ANS binding revealed that two lifetime components, 16.99 ns and 2.76 ns at pH 7.3, have dissociation constants of 0.58 mu M and 5.7 mu M, respectively. At pH 3.0, the lifetime components show decreased affinities with dissociation constants of 2.42 mu M and similar to 21 mu M, respectively. Selective displacement of ANS molecules from the ANS-apoTL complex by stearic acid discriminates the internal and external binding sites. Dependence of the binding affinity on ionic strength under various conditions provides strong evidence that an electrostatic interaction is involved. Time-resolved fluorescence is a promising tool to segregate multiple binding sites of proteins. (C) 2007 Elsevier Inc. All rights reserved.
作者
我是这篇论文的作者
点击您的名字以认领此论文并将其添加到您的个人资料中。
推荐
暂无数据