Comparative Study of the Gating Motif and C-type Inactivation in Prokaryotic Voltage-gated Sodium Channels*

Abstract

Prokaryotic voltage-gated sodium channels (Na_Vs) are homotetramers and are thought to inactivate through a single mechanism, named C-type inactivation. Here we report the voltage dependence and inactivation rate of the NaChBac channel from Bacillus halodurans, the first identified prokaryotic Na_V, as well as of three new homologues cloned from Bacillus licheniformis (Na_VBacL), Shewanella putrefaciens (Na_VSheP), and Roseobacter denitrificans (Na_VRosD). We found that, although activated by a lower membrane potential, Na_VBacL inactivates as slowly as NaChBac. Na_VSheP and Na_VRosD inactivate faster than NaChBac. Mutational analysis of helix S6 showed that residues corresponding to the “glycine hinge” and “PXP motif” in voltage-gated potassium channels are not obligatory for channel gating in these prokaryotic Na_Vs, but mutations in the regions changed the inactivation rates. Mutation of the region corresponding to the glycine hinge in Na_VBacL (A214G), Na_VSheP (A216G), and NaChBac (G219A) accelerated inactivation in these channels, whereas mutation of glycine to alanine in the lower part of helix S6 in NaChBac (G229A), Na_VBacL (G224A), and Na_VRosD (G217A) reduced the inactivation rate. These results imply that activation gating in prokaryotic Na_Vs does not require gating motifs and that the residues of helix S6 affect C-type inactivation rates in these channels.