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Little is known about the structure of the C terminus of the human cardiac voltage-gated sodium channel α subunit (SCN5A), but disease-linked mutations within this 244-amino acid intracellular region of the channel have marked effects on channel inactivation. Here we report a structural analysis of the C-terminal tail of the cardiac Na+channel that sheds new light on mechanisms that control its inactivation gating. Homology modeling of the SCN5AC terminus predicts predominant α-helical structure (six helices) in the proximal half of this intracellular tail but little structure in the distal half. Circular dichroism of isolated and purified C terminus supports this prediction. Whole cell and single channel patch clamp recordings of wild type and mutant α subunits co-expressed with the hβ1subunit in HEK 293 cells indicate that truncation of the distal, nonstructured, C terminus (L1921stop mutant) reduces current density but does not affect channel gating (n= 6). In contrast, truncation of the sixth helix containing a concentration of positively charged residues along with the distal C terminus (S1885stop mutant) also reduces current density but, in addition, has profound and selective effects on inactivation (no effect on activation). Channel availability is shifted (−11 ± 0.6 mV), and there is a 10-fold increase in the percentage of channels that burst (fail to inactivate) during prolonged depolarization (0.025% S1885stop (n= 7)versus0.0028% wild type (n= 9),p< 0.005). These results suggest that the charged structured region of the SCN5AC terminus plays a major role in channel inactivation, stabilizing the inactivated state.