Guanidinium chloride- and urea-induced unfolding of FprA, a mycobacterium NADPH-ferredoxin reductase - Stabilization of an apo-protein by GdmCl

被引:24
作者
Shukla, N
Bhatt, AN
Aliverti, A
Zanetti, G
Bhakuni, V [1 ]
机构
[1] Cent Drug Res Inst, Div Mol & Struct Biol, Lucknow 226001, Uttar Pradesh, India
[2] Univ Milan, Dipartimento Sci Biomol & Biotechnol, Milan, Italy
关键词
circular dichroism; electrostatic inteaction; fluorescence; FprA; chloride; intermediates;
D O I
10.1111/j.1742-4658.2005.2005.04645.x
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
The guanidinium chloride- and urea-induced unfolding of FprA, a mycobacterium NADPH-ferredoxin reductase, was examined in detail using multiple spectroscopic techniques, enzyme activity measurements and size exclusion chromatography. The equilibrium unfolding of FprA by urea is a cooperative process where no stabilization of any partially folded intermediate of protein is observed. In comparison, the unfolding of FprA by guanidinium chloride proceeds through intermediates that are stabilized by interaction of protein with guanidinium chloride. In the presence of low concentrations of guanidinium chloride the protein undergoes compaction of the native conformation; this is due to optimization of charge in the native protein caused by electrostatic shielding by the guanidinium cation of charges on the polar groups located on the protein side chains. At a guanidinium chloride concentration of about 0.8 M, stabilization of apo-protein was observed. The stabilization of apo-FprA by guanidinium chloride is probably the result of direct binding of the Gdm(+) cation to protein. The results presented here suggest that the difference between the urea- and guanidinium chloride-induced unfolding of FprA could be due to electrostatic interactions stabilizating the native conformation of this protein.
引用
收藏
页码:2216 / 2224
页数:9
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