We address the contribution of kinase domain structure and catalytic activity to membrane trafficking of TrkA receptor tyrosine kinase. We conduct a systematic comparison between TrkA-wt, an ATP-binding defective mutant (TrkA-K544N) and other mutants displaying separate functional impairments of phosphorylation, ubiquitination, or recruitment of intracellular partners. We find that only K544N mutation endows TrkA with restricted membrane mobility and a substantial increase of cell surface pool already in the absence of ligand stimulation. This mutation is predicted to drive a structural destabilization of the αC helix in the N-lobe by molecular dynamics simulations, and enhances interactions with elements of the actin cytoskeleton. On the other hand, a different TrkA membrane immobilization is selectively observed after NGF stimulation, requires both phosphorylation and ubiquitination to occur, and is most probably related to the signaling abilities displayed by the wt but not mutated receptors. In conclusion, our results allow to distinguish two different TrkA membrane immobilization modes and demonstrate that not all kinase-inactive mutants display identical membrane trafficking.

中文翻译：

---

TrkA激酶死亡突变体的膜运输改变的分子洞察力。

我们解决了激酶结构域结构和催化活性的膜运输的TrkA受体酪氨酸激酶的贡献。我们进行了TrkA-wt，ATP结合缺陷突变体（TrkA-K544N）与其他突变体之间的系统比较，这些突变体分别显示了磷酸化，泛素化或募集细胞内伴侣的功能受损。我们发现只有K544N突变赋予TrkA膜移动性受到限制，并且在没有配体刺激的情况下，细胞表面池大量增加。通过分子动力学模拟，预测该突变将驱动N叶中αC螺旋的结构失稳，并增强与肌动蛋白细胞骨架元素的相互作用。另一方面，NGF刺激后选择性观察到了不同的TrkA膜固定，需要同时发生磷酸化和泛素化，并且最可能与野生型显示的信号传导能力有关，但与突变受体无关。总之，我们的结果可以区分两种不同的TrkA膜固定模式，并证明并非所有激酶无活性的突变体都显示相同的膜运输。