Departments of Chemical Physiology and Molecular Biology, The Scripps Research Institute, La Jolla, CA 92037, USA.
Proc Natl Acad Sci U S A. 2011 Jul 26;108(30):12307-12. doi: 10.1073/pnas.1104293108. Epub 2011 Jul 7.
The question of how dispersed mutations in one protein engender the same gain-of-function phenotype is of great interest. Here we focus on mutations in glycyl-tRNA synthetase (GlyRS) that cause an axonal form of Charcot-Marie-Tooth (CMT) diseases, the most common hereditary peripheral neuropathies. Because the disease phenotype is dominant, and not correlated with defects in the role of GlyRS in protein synthesis, the mutant proteins are considered to be neomorphs that gain new functions from altered protein structure. Given that previous crystal structures showed little conformational difference between dimeric wild-type and CMT-causing mutant GlyRSs, the mutant proteins were investigated in solution by hydrogen-deuterium exchange (monitored by mass spectrometry) and small-angle X-ray scattering to uncover structural changes that could be suppressed by crystal packing interactions. Significantly, each of five spatially dispersed mutations induced the same conformational opening of a consensus area that is mostly buried in the wild-type protein. The identified neomorphic surface is thus a candidate for making CMT-associated pathological interactions, and a target for disease correction. Additional result showed that a helix-turn-helix WHEP domain that was appended to GlyRS in metazoans can regulate the neomorphic structural change, and that the gain of function of the CMT mutants might be due to the loss of function of the WHEP domain as a regulator. Overall, the results demonstrate how spatially dispersed and seemingly unrelated mutations can perpetrate the same localized effect on a protein.
一个蛋白质中分散突变如何产生相同的功能获得表型这一问题引起了广泛关注。在这里，我们重点研究甘氨酰-tRNA 合成酶（GlyRS）中的突变，这些突变会导致遗传性周围神经病中最常见的轴索型腓骨肌萎缩症（CMT）。由于疾病表型是显性的，并且与 GlyRS 在蛋白质合成中的作用缺陷无关，因此这些突变蛋白被认为是获得新功能的新形态，其结构发生了改变。鉴于之前的晶体结构显示二聚体野生型和引起 CMT 的突变 GlyRS 之间的构象差异很小，因此通过氢氘交换（通过质谱监测）和小角度 X 射线散射对突变蛋白在溶液中的结构进行了研究，以揭示可能被晶体包装相互作用抑制的结构变化。重要的是，五个空间分散的突变中的每一个都诱导了一个共识区域的相同构象开放，该区域在野生型蛋白中大部分被埋藏。因此，所鉴定的新形态表面是引起 CMT 相关病理相互作用的候选者，也是疾病矫正的目标。其他结果表明，后生动物中添加到 GlyRS 上的螺旋-环-螺旋 WHEP 结构域可以调节新形态的结构变化，并且 CMT 突变体的功能获得可能是由于 WHEP 结构域作为调节剂的功能丧失所致。总的来说，这些结果表明，空间分散且看似无关的突变如何对蛋白质产生相同的局部影响。
