Possible Roles of Exceptionally Conserved Residues around the Selectivity Filters of Sodium and Calcium Channels

Abstract

In the absence of x-ray structures of sodium and calcium channels their homology models are used to rationalize experimental data and design new experiments. A challenge is to model the outer-pore region that folds differently from potassium channels. Here we report a new model of the outer-pore region of the NaV1.4 channel, which suggests roles of highly conserved residues around the selectivity filter. The model takes from our previous study (Tikhonov, D. B., and Zhorov, B. S. (2005) Biophys. J. 88, 184 -197) the general disposition of the P-helices, selectivity filter residues, and the outer carboxylates, but proposes new intra-and inter-domain contacts that support structural stability of the outer pore. Glycine residues downstream from the selectivity filter are proposed to participate in knob-into-hole contacts with the P-helices and S6s. These contacts explain the adapted tetrodotoxin resistance of snakes that feed on toxic prey through valine substitution of isoleucine in the P-helix of repeat IV. Polar residues five positions upstream from the selectivity filter residues form H-bonds with the ascending-limb backbones. Exceptionally conserved tryptophans are engaged in inter-repeat H-bonds to form a ring whose -electrons would facilitate passage of ions from the outer carboxylates to the selectivity filter. The outerpore model of CaV1.2 derived from the NaV1.4 model is also stabilized by the ring of exceptionally conservative tryptophans and H-bonds between the P-helices and ascending limbs. In this model, the exceptionally conserved aspartate downstream from the selectivity-filter glutamate in repeat II facilitates passage of calcium ions to the selectivity-filter ring through the tryptophan ring. Available experimental data are discussed in view of the models.Voltage-gated ion channels are involved in the control of many physiological functions. Upon membrane depolarization, channels rapidly transit from the resting to the open state, then close to inactivated state(s), and return to the resting state upon membrane repolarization. The human genome encodes nine voltage-gated sodium (NaV1.1-NaV1.9) and ten voltage-gated calcium channels categorized as L (CaV1.1-CaV1.4), P/Q (CaV2.1), N (CaV2.2), R (CaV2.3), and T (CaV3.1-CaV3.3) types. The pore-forming ␣ 1 -subunit of calcium and sodium channels folds from a single polypeptide chain of four homologous repeats (Fig. 1A). Repeats I to IV are arranged clockwise when viewed extracellularly (1). Each repeat contains a voltage-sensing domain S1-S4, an outer helix S5, an inner helix S6, and a membrane-diving P-loop between S5 and S6. The P-loop includes a P-helix, a P-turn, and an ascending limb. Four ascending limbs, which line the outer pore, contain selectivity-filter residues in positions p50 (see Footnote 2 for residue labels). 2 Repeats I, II, III, and IV contain selectivity-filter glutamates in calcium channels (the EEEE ring) and Asp, Glu, Lys, and Ala residues in sodium channels (the DEKA ring). The ascending limbs also contain the outer carboxylates t...

Constraint shapes convergence in tetrodotoxin-resistant sodium channels of snakes

“…Nine of these substitutions replace amino acids in the β-strand, whose side chains face the pore and interact directly with TTX ( Fig. 2 and Movie S1) (35)(36)(37)(38). Mapping these changes and the TTX-resistance data onto the colubroid phylogeny (Fig.…”

“…This substitution occurs at a TTX-sensing residue (37), and the loss of a rigid alphatic side chain and gain of a larger functional group is expected to alter the orientation of the α-helix and thus the geometry of the outer pore, thereby indirectly weakening TTX-binding affinity to the pore (38).…”

“…We focused on the Ploops because TTX interacts with residues of the outer pore (see refs. [35][36][37][38], and changes in P-loop sites appear to be responsible for TTX resistance in animals (32,34,(39)(40)(41)(42)62).…”

“…We examined sequence variation in the four domains of Na v 1.4 (DI-DIV) that encode the outer pore of the channel (Ploops) and whose residues interact with TTX (35)(36)(37)(38) for six species of snakes known to prey on TTX-bearing amphibians, their sister groups, and additional taxa (74 species; n = 121 individuals) to provide a robust phylogenetic perspective (Methods). We scored organismal TTX resistance for nearly half these species (29 species; n = 424) using a performance bioassay (Methods).…”

Interaction of Tarantula Venom Peptide ProTx-II with Lipid Membranes Is a Prerequisite for Its Inhibition of Human Voltage-gated Sodium Channel NaV1.7

“…The molecular details of the inhibitory mechanism of pore blockers are well characterized. This has been facilitated by the extracellular location of the ion-conducting pore and by the overlapping binding sites of many pore blockers (12). Because the ion-conducting pore is highly conserved among channels of the same ion selectivity (e.g.…”