Sivakolundu Sivashankar G, Bashford Donald, Kriwacki Richard W
Department of Structural Biology, St. Jude Children's Research Hospital, 332 North Lauderdale St., Memphis, TN 38105, USA.
J Mol Biol. 2005 Nov 11;353(5):1118-28. doi: 10.1016/j.jmb.2005.08.074. Epub 2005 Sep 20.
p27Kip1 (p27) influences cell division by regulating nuclear cyclin-dependent kinases. Before binding, p27 is at least partially disordered and folds upon binding its Cdk/cyclin targets. 30-40% of human proteins, including p27, are predicted to contain disordered segments, and have been termed intrinsically unstructured proteins (IUPs). Unfortunately, the inherent dynamics of IUPs hamper detailed analysis of their structure/function relationships. Here, we describe the use of molecular dynamics (MD) computations and solution NMR spectroscopy to reveal that several segments of the p27 kinase inhibitory domain (p27-KID), in addition to the previously characterized helical segment, exist as highly populated, intrinsically folded structural units (IFSUs). Several IFSUs resemble structural features of bound p27-KID, while another exhibits alternative conformations. Interestingly, the highly conserved, specificity determining segment of p27 is shown to be highly disordered. Elucidation of IFSUs within p27-KID allows consideration of their influences on the thermodynamics and kinetics of Cdk/cyclin binding. The degree to which IFSUs are populated within p27-KID is surprising and suggests that other putative IUPs contain IFSUs that may be studied using similar techniques.
p27Kip1（p27）通过调节细胞核周期蛋白依赖性激酶来影响细胞分裂。在结合之前，p27至少部分处于无序状态，并在结合其细胞周期蛋白依赖性激酶/细胞周期蛋白靶点时发生折叠。包括p27在内的30%至40%的人类蛋白质预计含有无序片段，这些蛋白质被称为内在无序蛋白（IUP）。不幸的是，IUP的固有动力学阻碍了对其结构/功能关系的详细分析。在这里，我们描述了使用分子动力学（MD）计算和溶液核磁共振波谱来揭示，除了先前已表征的螺旋片段外，p27激酶抑制结构域（p27-KID）的几个片段以高度富集的内在折叠结构单元（IFSU）形式存在。几个IFSU类似于结合态p27-KID的结构特征，而另一个则呈现出不同的构象。有趣的是，p27高度保守的、决定特异性的片段被证明是高度无序的。阐明p27-KID内的IFSU有助于考虑它们对细胞周期蛋白依赖性激酶/细胞周期蛋白结合的热力学和动力学的影响。p27-KID内IFSU的富集程度令人惊讶，并表明其他假定的IUP含有可以使用类似技术进行研究的IFSU。
