Piconewton-Scale Analysis of Ras-BRaf Signal Transduction with Single-Molecule Force Spectroscopy.
Piconewton-Scale Analysis of Ras-BRaf Signal Transduction with Single-Molecule Force Spectroscopy.
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使用单分子力谱对 Ras-BRaf 信号转导进行皮牛顿尺度分析。
DOI：
10.1002/smll.201701972
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发表时间：
2017
期刊：
影响因子：
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通讯作者：
Zhu,JJulius
中科院分区：
文献类型：
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作者：
Lim,Chae-Seok;Wen,Cheng;Sheng,Yanghui;Wang,Guangfu;Zhou,Zhuan;Wang,Shiqiang;Zhang,Huaye;Ye,Anpei;Zhu,JJulius
分子间相互作用在细胞的各种生理和病理过程中主导着信号转导的行为，然而评估这些相互作用仍然是一个具有挑战性的任务。在这里，这项研究报告了一种单分子力谱方法，它能够在功能上描述典型的丝裂原活化蛋白激酶(MAPK)途径中信号效应器RAS和BRAF之间的两个相互作用位点(≈35PN和≈90PN)。这项分析揭示了BRAF上Q257和A246的突变，这两个经常与心面部皮肤综合征相关的位置导致了RAS≈10−30 Pn的RASBRAF分子间结合力的变化。RAS BRAF结合力变化的大小与表达替代BRAF突变体的神经元中蛋白质亲和力和α‐amino‐3‐hydroxy‐5‐methyl‐4‐isoxazolepropionic酸(AMPA)敏感的谷氨酸受体(-R)介导的突触传递的变化的大小相关，并预测表达替代突变体的动物学习障碍的程度。这些结果建立了单分子力谱作为一个有效的平台来评估信号分子之间的微微子水平的相互作用和预测信号转导的行为结果。
Intermolecular interactions dominate the behavior of signal transduction in various physiological and pathological cell processes, yet assessing these interactions remains a challenging task. Here, this study reports a single‐molecule force spectroscopic method that enables functional delineation of two interaction sites (≈35 pN and ≈90 pN) between signaling effectors Ras and BRaf in the canonical mitogen‐activated protein kinase (MAPK) pathway. This analysis reveals mutations on BRaf at Q257 and A246, two sites frequently linked to cardio‐faciocutaneous syndrome, result in ≈10−30 pN alterations in RasBRaf intermolecular binding force. The magnitude of changes in RasBRaf binding force correlates with the size of alterations in protein affinity and in α‐amino‐3‐hydroxy‐5‐methyl‐4‐isoxazolepropionic acid (AMPA)‐sensitive glutamate receptor (‐R)‐mediated synaptic transmission in neurons expressing replacement BRaf mutants, and predicts the extent of learning impairments in animals expressing replacement BRaf mutants. These results establish single‐molecule force spectroscopy as an effective platform for evaluating the piconewton‐level interaction of signaling molecules and predicting the behavior outcome of signal transduction.