Laboratory of Membrane Biology and Biophysics, The Rockefeller University, New York, NY 10065.
Department of Biochemistry, Semmelweis University, Budapest H-1094, Hungary.
Proc Natl Acad Sci U S A. 2024 Nov 12;121(46):e2409049121. doi: 10.1073/pnas.2409049121. Epub 2024 Nov 4.
Protein kinase A (PKA) is a key regulator of cellular functions by selectively phosphorylating numerous substrates, including ion channels, enzymes, and transcription factors. It has long served as a model system for understanding the eukaryotic kinases. Using cryoelectron microscopy, we present complex structures of the PKA catalytic subunit (PKA-C) bound to a full-length protein substrate, the cystic fibrosis transmembrane conductance regulator (CFTR)-an ion channel vital to human health. CFTR gating requires phosphorylation of its regulatory (R) domain. Unphosphorylated CFTR engages PKA-C at two locations, establishing two "catalytic stations" near to, but not directly involving, the R domain. This configuration, coupled with the conformational flexibility of the R domain, permits transient interactions of the eleven spatially separated phosphorylation sites. Furthermore, we determined two structures of the open-pore CFTR stabilized by PKA-C, providing a molecular basis to understand how PKA-C stimulates CFTR currents even in the absence of phosphorylation.
蛋白激酶 A（PKA）通过选择性磷酸化众多底物，包括离子通道、酶和转录因子，来调节细胞功能，它长期以来一直是理解真核激酶的模型系统。我们使用冷冻电子显微镜，呈现了 PKA 催化亚基（PKA-C）与全长蛋白底物囊性纤维化跨膜电导调节剂（CFTR）结合的复合物结构，CFTR 是对人类健康至关重要的离子通道。CFTR 的门控需要其调节（R）域的磷酸化。未磷酸化的 CFTR 在两个位置与 PKA-C 结合，在 R 域附近建立两个“催化站”，但不直接涉及 R 域。这种构象，加上 R 域的构象灵活性，允许十一个空间分离的磷酸化位点的瞬时相互作用。此外，我们还确定了两个由 PKA-C 稳定的开放孔 CFTR 结构，为理解 PKA-C 如何在没有磷酸化的情况下刺激 CFTR 电流提供了分子基础。
