Cregut D, Serrano L
European Molecular Biology Laboratory, Heidelberg, Germany.
Protein Sci. 1999 Feb;8(2):271-82. doi: 10.1110/ps.8.2.271.
The usefulness of molecular dynamics to assess the structural integrity of mutants containing several mutations has been investigated. Our goal was to determine whether molecular dynamics would be able to discriminate mutants of a protein having a close-to-wild-type fold, from those that are not folded under the same conditions. We used as a model the B1 domain of protein G in which we replaced the unique central alpha-helix by the sequence of the second beta-hairpin, which has a strong intrinsic propensity to form this secondary structure in solution. In the resulting protein, one-third of the secondary structure has been replaced by a non-native one. Models of the mutants were built based on the three-dimensional structure of the wild-type GB1 domain. During 2 ns of molecular dynamics simulations on these models, mutants containing up to 10 mutations in the helix retained the native fold, while another mutant with an additional mutation unfolded. This result is in agreement with our circular dichroism and NMR experiments, which indicated that the former mutants fold into a structure similar to the wild-type, as opposed to the latter mutant which is partly unfolded. Additionally, a mutant containing six mutations scattered through the surface of the domain, and which is unfolded, was also detected by the simulation. This study suggests that molecular dynamics calculations could be performed on molecular models of mutants of a protein to evaluate their foldability, prior to a mutagenesis experiment.
已对分子动力学评估含有多个突变的突变体结构完整性的有用性进行了研究。我们的目标是确定分子动力学是否能够区分具有接近野生型折叠的蛋白质突变体与在相同条件下未折叠的突变体。我们使用蛋白G的B1结构域作为模型，在该模型中，我们用第二个β-发夹的序列替换了唯一的中央α-螺旋，该β-发夹在溶液中具有很强的形成这种二级结构的内在倾向。在所得蛋白质中，三分之一的二级结构已被非天然结构所取代。基于野生型GB1结构域的三维结构构建了突变体模型。在对这些模型进行2纳秒的分子动力学模拟过程中，螺旋中含有多达10个突变的突变体保留了天然折叠，而另一个带有额外突变的突变体则发生了去折叠。这一结果与我们的圆二色性和核磁共振实验结果一致，这些实验表明，前者的突变体折叠成与野生型相似的结构，而后者的突变体则部分去折叠。此外，模拟还检测到一个含有六个分散在结构域表面的突变且未折叠的突变体。这项研究表明，在诱变实验之前，可以对蛋白质突变体的分子模型进行分子动力学计算，以评估其可折叠性。
