The quaternary structure of streptavidin in urea.
The quaternary structure of streptavidin in urea.
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DOI：
10.1016/s0021-9258(18)98710-7
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发表时间：
1991-08
期刊：
影响因子：
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通讯作者：
G. Kurzban;Edward A. Bayer;M. Wilchek;Paul M. Horowitz
中科院分区：
文献类型：
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作者：
G. Kurzban;Edward A. Bayer;M. Wilchek;Paul M. Horowitz
我们报道了脲和胍盐与链霉亲和素的相互作用。在0、4、6和7 M尿素中凝胶过滤层析表明链霉亲和素四聚体在尿素中保持完整。无论是否存在6m尿素，生物素都会改变链霉亲和素的电泳迁移率。链霉亲和素的本征荧光在6 M尿素中增强和蓝移。荧光变化表明没有展开。对尿素的构象响应是可能的，但大部分荧光变化是由于尿素作为弱生物素类似物结合（Ka约为1.3 M-1）。抗尿素结构扰动反映了链霉亲和素抗平行β -桶基序的结构稳定性。在6 M盐酸胍中展开缓慢（半衰期，约50天）。硫氰酸胍展开后，链亲和素可以再折叠，但展开和再折叠的过渡集中在不同浓度的扰动上。缓慢展开，以15次方依赖于硫氰酸胍浓度，可能部分负责展开和再折叠过程的不重合。在6 M尿素中也可以看到非平衡行为，因为天然链霉亲和素不展开，而硫氰酸胍不展开链霉亲和素不折叠。在较低浓度的尿素中确实会发生再折叠。硫氰酸胍只能缓慢地展开生物素-链亲和素复合物。在生物素存在的情况下，未折叠的链亲和素在6m硫氰酸胍或6m尿素中不会再折叠。
We report on the interactions of urea and guanidinium salts with streptavidin. Gel filtration chromatography in 0, 4, 6, and 7 M urea indicates that the streptavidin tetramer remains intact in urea. Biotin alters the electrophoretic mobility of streptavidin whether or not 6 M urea is present. The intrinsic fluorescence of streptavidin is increased and blue-shifted in 6 M urea. The fluorescence changes indicate the absence of unfolding. A conformational response to urea is possible, but much of the fluorescence change is due to urea binding as a weak biotin analog (Ka approximately 1.3 M-1). The resistance to structural perturbation by urea reflects the structural stability of streptavidin's anti-parallel beta-barrel motif. Unfolding is sluggish in 6 M guanidinium hydrochloride (half-time, approximately 50 days). After guanidinium thiocyanate unfolding, streptavidin can be refolded, but the unfolding and refolding transitions are centered at different concentrations of perturbant. Slow unfolding, with a 15th power dependence on guanidinium thiocyanate concentration, may be partially responsible for the noncoincidence of the unfolding and refolding processes. Nonequilibrium behavior is also seen in 6 M urea, as native streptavidin does not unfold and guanidinium thiocyanate unfolded streptavidin does not refold. Refolding does occur at lower concentrations of urea. Guanidinium thiocyanate only slowly unfolds the biotin-streptavidin complex. In the presence of biotin, unfolded streptavidin does not refold in 6 M guanidinium thiocyanate or in 6 M urea.