Department of Materials Science and Engineering, Johns Hopkins University, Baltimore, Maryland; Institute for NanoBioTechnology, Johns Hopkins University, Baltimore, Maryland.
Program in Molecular Biophysics, Johns Hopkins University, Baltimore, Maryland; Institute for NanoBioTechnology, Johns Hopkins University, Baltimore, Maryland.
Biophys J. 2021 Jan 5;120(1):55-63. doi: 10.1016/j.bpj.2020.11.2262. Epub 2020 Dec 5.
Trk-A is a receptor tyrosine kinase (RTK) that plays an essential role in the development and functioning of the nervous system. Trk-A is expressed in neurons and signals in response to two ligands, NGF and neurotrophin-3 (NT-3), with very different functional consequences. Thus, NGF and NT-3 are "biased" ligands for Trk-A. Because it has been hypothesized that biased RTK ligands induce differential stabilization of RTK dimers, here, we seek to test this hypothesis for NGF and NT-3. In particular, we use Förster resonance energy transfer (FRET) and fluorescence intensity fluctuation spectroscopy to assess the strength of Trk-A interactions and Trk-A oligomer size in the presence of the two ligands. Although the difference in Trk-A behavior in response to the two ligands has been previously attributed to differences in their binding to Trk-A in the endosomes at low pH, here, we further show differences in the stabilities of the NGF- and NT-3-bound Trk-A dimers in the plasma membrane and at neutral pH. We discuss the biological significance of these new findings and their implications for the design of Trk-A ligands with novel functionalities.
Trk-A 是一种受体酪氨酸激酶（RTK），在神经系统的发育和功能中起着至关重要的作用。Trk-A 在神经元中表达，并响应两种配体，神经生长因子（NGF）和神经营养因子-3（NT-3），而产生非常不同的功能后果。因此，NGF 和 NT-3 是 Trk-A 的“偏向”配体。因为有人假设偏向性 RTK 配体诱导 RTK 二聚体的不同稳定性，在这里，我们试图针对 NGF 和 NT-3 来检验这一假设。具体来说，我们使用荧光共振能量转移（FRET）和荧光强度波动光谱来评估两种配体存在时 Trk-A 相互作用的强度和 Trk-A 寡聚体的大小。尽管先前已经将 Trk-A 对两种配体的反应差异归因于它们在低 pH 时在内体中与 Trk-A 的结合差异，但在这里，我们进一步显示了在质膜中和中性 pH 下，NGF 和 NT-3 结合的 Trk-A 二聚体的稳定性存在差异。我们讨论了这些新发现的生物学意义及其对具有新型功能的 Trk-A 配体设计的影响。
