Wijesinha-Bettoni R, Dobson C M, Redfield C
Oxford Centre for Molecular Sciences, Central Chemistry Laboratory, University of Oxford, South Parks Road, Oxford OX1 3QH, UK.
J Mol Biol. 2001 Sep 7;312(1):261-73. doi: 10.1006/jmbi.2001.4927.
Residue-specific information on the urea-induced unfolding of the molten globule state of bovine alpha-lactalbumin (BLA) has been obtained using NMR spectroscopy. In agreement with previous studies on human alpha-lactalbumin (HLA) the unfolding process for BLA has been found to be non-cooperative. Both the alpha and beta-domains of the protein are substantially collapsed in the absence of denaturant but in both proteins the majority of the structure in the beta-domain unfolds prior to that in the alpha-domain. However, in BLA the protein unfolds completely in 10 M urea at 50 degrees C, whilst in HLA a stable core region persists even under these extreme conditions. Previous studies on HLA have identified eight residues that are crucial for the stability of the molten globule. Of these residues, only three are conserved in the sequence of BLA. By taking into consideration the differences in inter-residue contacts between the four alpha-helices arising from these substitutions, and the relative hydrophobicity of the helices in the two proteins, we show that it is possible to rationalise the observed differences in the behaviour of the molten globule states of the two proteins. Taken together, these results suggest that there may be a number of ways of stabilising a given protein fold, and the specific manner that this is achieved for a particular protein is determined by details of its sequence.
利用核磁共振光谱法已获得了关于尿素诱导牛α-乳白蛋白（BLA）熔融球状态展开的残基特异性信息。与之前对人α-乳白蛋白（HLA）的研究一致，发现BLA的展开过程是非协同的。在没有变性剂的情况下，该蛋白质的α结构域和β结构域都基本折叠，但在这两种蛋白质中，β结构域中的大部分结构在α结构域之前就展开了。然而，在50℃下，BLA在10M尿素中能完全展开，而在HLA中，即使在这些极端条件下，仍有一个稳定的核心区域存在。之前对HLA的研究已经确定了八个对熔融球状态稳定性至关重要的残基。在这些残基中，只有三个在BLA序列中保守。通过考虑这些取代导致的四个α螺旋之间残基间接触的差异，以及两种蛋白质中螺旋的相对疏水性，我们表明有可能对观察到的两种蛋白质熔融球状态行为的差异做出合理的解释。综上所述，这些结果表明，稳定特定蛋白质折叠可能有多种方式，而特定蛋白质实现这一目标的具体方式取决于其序列细节。
