標題: Rational Design for Crystallization of beta-Lactoglobulin and Vitamin D(3) Complex: Revealing a Secondary Binding Site
作者: Yang, Ming Chi
Guan, Hong-Hsiang
Yang, Jirin-Moon
Ko, Cheng-Neng
Liu, Ming-Yih
Lin, Yih-Hung
Huang, Yen-Chieh
Chen, Chun-Jung
Mao, Simon J. T.
生物科技學系
Department of Biological Science and Technology
公開日期: 1-Dec-2008
摘要: beta-Lactoglobulin (LG) is a major milk whey protein containing primarily a calyx for vitamin D(3) binding, although the existence of another site beyond the calyx is controversial. Using fluorescence spectral analyses in the previous study, we showed the binding stoichiometry for vitamin D3 to LG to be 2:1 and a stoichiometry of 1:1 when the calyx was "disrupted" by manipulating the pH and temperature, suggesting that a secondary vitamin D binding site existed. To help localize this secondary site using X-ray crystallography in the present study, we used bioinformatic programs (Insight II, Q-SiteFinder, and GEMDOCK) to identify the potential location of this site. We then optimized the occupancy and enhanced the electron density of vitamin D3 in the complex by altering the pH and initial ratios of vitamin D(3)/LG in the cocrystal preparation. We conclude that GEMDOCK can aid in searching for an extra density map around potential vitamin D binding sites. Both pH (8) and initial ratio of vitamin D(3)/LG (3:1) are crucial to optimize the occupancy and enhance the electron density of vitamin D(3) in the complex for rational-designed crystallization. The strategy in practice may be useful for future identification of a ligand-binding site in a given protein.
URI: http://dx.doi.org/10.1021/cg800697s
http://hdl.handle.net/11536/28621
ISSN: 1528-7483
DOI: 10.1021/cg800697s
期刊: CRYSTAL GROWTH & DESIGN
Volume: 8
Issue: 12
起始頁: 4268
結束頁: 4276
Appears in Collections:Conferences Paper


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