Structural determination of human Nav1.4 and Nav1.7 using single particle cryo-electron microscopy

Abstract

Voltage-gated sodium (Na_v) channels are responsible for the initiation and propagation of action potentials. Their abnormal functions are associated with numerous diseases, such as epilepsy, cardiac arrhythmia, and pain syndromes. Therefore, these channels represent important drug targets. Even in the post-resolution revolution era, a lack of structural information continues to impede structure-based drug discovery. The limiting factor for the structural determination of Na_v channels using single particle cryo-electron microscopy (cryo-EM) resides in the generation of sufficient high-quality recombinant proteins. After extensive trials, we have been successful in determining a series of high-resolution structures of Na_v channels, including Na_vPaS from American cockroach, Na_v1.4 from electric eel, and human Na_v1.1, Na_v1.2, Na_v1.4, Na_v1.5, and Na_v1.7, with distinct strategies. These structures established the framework for understanding the electromechanical coupling and disease mechanism of Na_v channels, and for facilitating drug discovery. Here, we exemplify these methods with two specific cases, human Na_v1.4 and Na_v1.7, which may shed light on the structural determination of other membrane proteins.

Keywords: BacMam; Detergent; Na(v)1.4; Na(v)1.7; Protein optimization; Recombinant expression; Single particle cryo-EM.