Rogers Joseph M, Wong Chi T, Clarke Jane
Department of Chemistry, University of Cambridge , Lensfield Road, Cambridge, CB2 1EW, United Kingdom.
J Am Chem Soc. 2014 Apr 9;136(14):5197-200. doi: 10.1021/ja4125065. Epub 2014 Mar 31.
Many cellular proteins are 'disordered' in isolation. A subset of these intrinsically disordered proteins (IDPs) can, upon binding another molecule, fold to a well-defined three-dimensional structure. In the structurally heterogeneous, unbound ensemble of these IDPs, conformations are likely to exist that, in part, resemble the final bound form. It has been suggested that these conformations, displaying 'residual structure', could be important for the mechanism of such coupled folding and binding reactions. PUMA, of the BCL-2 family, is an IDP in isolation but will form a single, contiguous α-helix upon binding the folded protein MCL-1. Using the helix-breaking residue proline, we systematically target each potential turn of helix of unbound PUMA and assess the binding to MCL-1 using time-resolved stopped-flow techniques. All proline-containing mutants bound, and although binding was weaker than the wild-type protein, association rate constants were largely unaffected. We conclude that population of particular residual structure, containing a specific helical turn, is neither required for the binding nor for fast association of PUMA and MCL-1.
许多细胞蛋白单独存在时是“无序的”。这些内在无序蛋白（IDP）中的一部分在与另一个分子结合后，能够折叠成明确的三维结构。在这些IDP结构异质的未结合聚集体中，很可能存在部分类似于最终结合形式的构象。有人提出，这些呈现“残余结构”的构象对于此类偶联折叠和结合反应的机制可能很重要。BCL - 2家族的PUMA单独存在时是一种IDP，但在与折叠蛋白MCL - 1结合时会形成单一的连续α螺旋。我们使用螺旋破坏残基脯氨酸，系统地靶向未结合的PUMA螺旋的每个潜在转角，并使用时间分辨停流技术评估其与MCL - 1的结合。所有含脯氨酸的突变体都能结合，尽管结合力比野生型蛋白弱，但缔合速率常数基本不受影响。我们得出结论，含有特定螺旋转角的特定残余结构群体对于PUMA和MCL - 1的结合及快速缔合既不是必需的。
