Abstract
Voltage-gated sodium channels play essential roles in many important physiological processes, including electric signalling and muscle contraction. Therefore their activation and inactivation are needed to be strictly regulated.Here, we show that the cytosolic C-terminal region of NavSulP, a prokaryotic voltage-gated sodium channel (NavBac) cloned from Sulfitobacter pontiacus, accelerates C-type inactivation. The crystal structure of the C-terminal region of NavSulP in a NaK-NavSulP chimera channel at 3.2 A resolution revealed that the C-terminal region forms a four-helix bundle. Point mutations of the residues of the four-helix bundle, which were involved in intersubunit interactions, destabilised the tetramer of the channel and reduced the inactivation rate. This result suggested that the formation of the four-helix bundle accelerates the inactivation of NavSulP. NavBac inactivation seems to be regulated only by the C-type inactivation, which is thought to be due to collapse of the selectivity filter through a conformational change of the activation gate of the inner helix of the pore domain. The C-terminal four-helix bundle is connected to the inner helix. The increase of the rigidity of the inner helix with the glycine-to-alanine mutation also reduced the inactivation rate of NavSulP as well as the destabilisation of the four-helix bundle. On the other hand, the glycine-to-alanine mutation did not destabilize the formation of tetrameric channel, and then the C-terminal region could maintain the four-helix bundle. It was thought that the rigidity of inner helix disabled the formation of C-terminal four-helix bundle from accelerating the C-type inactivation rate.These findings suggest that the formation of four-helix bundle of NavSulP plays important roles not only in stabilising the tetramer, but also in accelerating C-type inactivation by promoting a conformational change of the inner helices of the pore domain.
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