Switching of the folding-energy landscape governs the allosteric activation of protein kinase A
Switching of the folding-energy landscape governs the allosteric activation of protein kinase A
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DOI：
10.1073/pnas.1802510115
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发表时间：
2018-08-07
影响因子：
11.1
通讯作者：
Maillard, Rodrigo A.
中科院分区：
文献类型：
--
作者：
England, Jeneffer P.;Hao, Yuxin;Maillard, Rodrigo A.
蛋白激酶是对多种构象状态进行采样的动态分子开关。PKA的调节亚基含有两个cAMP结合结构域[环核苷酸结合（CNB）结构域]，它们依赖于cAMP结合而在非活性和活性构象之间振荡。cAMP与CNB结构域的协同结合激活了变构相互作用网络，使PKA能够从非活性构象发展为活性构象，释放催化亚基的活性。尽管它在许多生物过程的调节中的重要性，PKA的激活过程中观察到的协同性的分子机制仍然不清楚。在这里，我们使用光镊探测的折叠协同性和能量的cAMP结合域之间的通信在载脂蛋白状态和绑定到催化亚基。我们的研究提供了直接的证据，在折叠能量景观的两个CNB域的开关，从能量独立的载脂蛋白状态高度合作，并在催化亚基的存在下大力耦合。此外，我们表明，在一个CNB结构域的不稳定突变的影响有效地传播到其他和降低它们之间的折叠协同性。总之，我们的研究结果提供了一个热力学基础的构象可塑性，使蛋白激酶适应和响应信号分子。
Protein kinases are dynamic molecular switches that sample multiple conformational states. The regulatory subunit of PKA harbors two cAMP-binding domains [cyclic nucleotide-binding (CNB) domains] that oscillate between inactive and active conformations dependent on cAMP binding. The cooperative binding of cAMP to the CNB domains activates an allosteric interaction network that enables PKA to progress from the inactive to active conformation, unleashing the activity of the catalytic subunit. Despite its importance in the regulation of many biological processes, the molecular mechanism responsible for the observed cooperativity during the activation of PKA remains unclear. Here, we use optical tweezers to probe the folding cooperativity and energetics of domain communication between the cAMP-binding domains in the apo state and bound to the catalytic subunit. Our study provides direct evidence of a switch in the folding-energy landscape of the two CNB domains from energetically independent in the apo state to highly cooperative and energetically coupled in the presence of the catalytic subunit. Moreover, we show that destabilizing mutational effects in one CNB domain efficiently propagate to the other and decrease the folding cooperativity between them. Taken together, our results provide a thermodynamic foundation for the conformational plasticity that enables protein kinases to adapt and respond to signaling molecules.