Allosteric coupling asymmetry mediates paradoxical activation of BRAF by type II inhibitors.
Allosteric coupling asymmetry mediates paradoxical activation of BRAF by type II inhibitors.
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变构偶联不对称介导 II 型抑制剂对 BRAF 的反常激活。
DOI：
10.1101/2023.04.18.536450
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发表时间：
2023
期刊：
影响因子：
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通讯作者：
Levinson,NicholasM
中科院分区：
文献类型：
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作者：
Rasmussen,DamienM;Semonis,MannyM;Greene,JosephT;Muretta,JosephM;Thompson,AndrewR;Ramos,SilviaToledo;Thomas,DavidD;Pomerantz,WilliamCK;Freedman,TanyaS;Levinson,NicholasM
目前在临床试验中的II型RAF抑制剂在亚饱和浓度下矛盾地激活BRAF。激活是通过诱导BRAF二聚体介导的，但为什么激活而不是抑制发生仍然不清楚。使用跟踪BRAF二聚化和构象的生物物理方法，我们建立了一个变构模型，解决了抑制剂结合到二聚体的两个活性位点的变构贡献，揭示了I型和II型RAF抑制剂之间的关键差异。对于II型抑制剂，抑制剂结合和BRAF二聚化之间的变构偶联不对称地分布在两个二聚体结合位点上，与第一个位点的结合主导变构。这种不对称性导致有效和选择性诱导二聚体与一个抑制和一个催化活性亚基。我们的变构模型定量解释了11种RAF抑制剂测量的反常激活数据。与II型抑制剂不同，I型抑制剂缺乏变构不对称性并且不激活BRAF同二聚体。最后，NMR数据显示BRAF同源二聚体是动态不对称的，只有一个亚基锁定在活性αC-in状态。这为仅结合单个αC-in抑制剂分子如何诱导有效的BRAF二聚化和活化提供了结构机制。
The type II class of RAF inhibitors currently in clinical trials paradoxically activate BRAF at subsaturating concentrations. Activation is mediated by induction of BRAF dimers, but why activation rather than inhibition occurs remains unclear. Using biophysical methods tracking BRAF dimerization and conformation, we built an allosteric model of inhibitor-induced dimerization that resolves the allosteric contributions of inhibitor binding to the two active sites of the dimer, revealing key differences between type I and type II RAF inhibitors. For type II inhibitors the allosteric coupling between inhibitor binding and BRAF dimerization is distributed asymmetrically across the two dimer binding sites, with binding to the first site dominating the allostery. This asymmetry results in efficient and selective induction of dimers with one inhibited and one catalytically active subunit. Our allosteric models quantitatively account for paradoxical activation data measured for 11 RAF inhibitors. Unlike type II inhibitors, type I inhibitors lack allosteric asymmetry and do not activate BRAF homodimers. Finally, NMR data reveal that BRAF homodimers are dynamically asymmetric with only one of the subunits locked in the active αC-in state. This provides a structural mechanism for how binding of only a single αC-in inhibitor molecule can induce potent BRAF dimerization and activation.