Control of Slow, Use Dependent Inactivation of NaVAb by its C Terminal Tail

Abstract

Bacterial voltage-gated sodium channels are composed of four identical subunits. They share major biophysical features with eukaryotic counterparts. Crystallization of full-length bacterial sodium channels (Payandeh et al. 2011, 2012; Zhang et al. 2012) makes them invaluable models for studying the structural basis of ion conductance, activation, inactivation, and drug interaction. Mammalian sodium channels have two main types of inactivation: fast inactivation on the order of milliseconds, which is mediated by the IFM motif in the intracellular loop between domains III and IV; and slow inactivation on the order of hundreds of milliseconds to seconds, which is believed to occur through pore collapse. The bacterial sodium channel NavAb activates at very negative membrane potentials, and it has a late use-dependent phase of slow inactivation that reverses very slowly (Gamal El-Din et al. 2013). We studied the effect of the NavAb C-terminal cytoplasmic domain on use-dependent slow inactivation. Deletion of 40 residues of the cytoplasmic tail (α40) abolished late use-dependent inactivation. Progressively smaller deletions to yield α28, α10, α7, and α3 caused graded effects. However, deletion of only 10 residues was sufficient to abolish most of the late, use-dependent inactivation of NavAb. In addition to modulating the extent of use-dependent inactivation, the progressively truncated constructs showed decremental slowing of the decay of sodium current during depolarizations. NaVAb-α40 has voltage-independent kinetics of current decay, while other constructs have differential profiles of decay kinetics that are correlated with C-terminal length. Our experiments reveal a surprisingly crucial role for the C-terminal domain, which forms a coiled-coil structure unique to bacterial sodium channels, in both early and late phases of use-dependent slow inactivation of NaVAb.