Taylor L S, York P, Williams A C, Edwards H G, Mehta V, Jackson G S, Badcoe I G, Clarke A R
Postgraduate Studies in Pharmaceutical Technology, School of Pharmacy, University of Bradford, UK.
Biochim Biophys Acta. 1995 Nov 15;1253(1):39-46. doi: 10.1016/0167-4838(95)00142-h.
Sugars and polyols are used to stabilize proteins. The degree of stabilization conferred on a model protein by sucrose was calculated in terms of the free energy of folding. Phosphoglycerate kinase (PGK) was denatured by guanidine hydrochloride (GuHCl) in different sucrose concentrations. The linear extrapolation method [1,2] was used to calculate the free energy of folding in the absence of denaturant. Although sucrose increased the concentration of GuHCl required to unfold the protein, the free energy of folding in water was unchanged. In order to probe the nature of the stabilizing effect of sucrose, an FT-Raman spectroscopic study of denaturant-polyol systems was undertaken. Investigations of interactions between GuHCl, urea or formamide and polyhydric compounds, revealed no evidence for hydrogen bonding or dipole-dipole associations. Polyhydric compounds caused minor changes in denaturant spectra although the converse was not observed. The structure of deuterated water changed on addition of denaturants. For non-ionic denaturants, addition of polyhydric solutes countered this change in water structure. Thus polyhydric compounds oppose the effect of denaturants on water structure. The observed increase in GuHCl concentration required to unfold PGK in the presence of sucrose may be attributed to this property of sucrose.
糖类和多元醇被用于稳定蛋白质。根据折叠自由能计算了蔗糖赋予模型蛋白的稳定程度。在不同蔗糖浓度下，用盐酸胍（GuHCl）使磷酸甘油酸激酶（PGK）变性。采用线性外推法[1,2]计算无变性剂时的折叠自由能。尽管蔗糖提高了使蛋白质展开所需的GuHCl浓度，但水中的折叠自由能并未改变。为了探究蔗糖稳定作用的本质，对变性剂 - 多元醇体系进行了傅里叶变换拉曼光谱研究。对GuHCl、尿素或甲酰胺与多元化合物之间相互作用的研究，未发现氢键或偶极 - 偶极缔合的证据。多元化合物使变性剂光谱发生了微小变化，不过未观察到相反情况。加入变性剂后，重水的结构发生了变化。对于非离子变性剂，加入多元溶质可抵消水结构的这种变化。因此，多元化合物对抗变性剂对水结构的影响。在蔗糖存在下使PGK展开所需的GuHCl浓度增加，可能归因于蔗糖的这一特性。
