Voltage-gated sodium (Nav) channels initiate action potentials in neurons and other excitable cells. Mutations in them cause inherited epilepsy, migraine, chronic pain, and periodic paralysis, and they are important molecular targets for drugs. New insights into the structure and function of Nav channels have come from our high-resolution x-ray crystallography studies of their bacterial ancestor NavAb. Here we propose to further define the structural basis for the key functional properties of mammalian Nav channels by building their characteristic structural features into NavAb and analyzing their high-resolution structure and function. (i) We will define the structural basis for activation of mammalian voltage sensors through studies of NavAb chimeras in which one or more voltage sensors are substituted with mammalian counterparts. Preliminary experiments show that a human Nav1.7 voltage sensor grafted onto NavAb induces the voltage-dependent gating properties of the Nav1.7 channel. The amino acid residues that define Nav1.7 voltage dependence of gating will be defined, the structures of resting and activated Nav channels will be determined, and the role of the voltage sensor in determining concerted versus sequential gating of the pore will be analyzed. (ii) We will probe the structural basis for ion selectivity in mammalian Nav channels through structural and functional studies of NavAb chimeras. Although NavAb is Na-selective, its detailed permeation properties for other monovalent ions are very different from mammalian Nav channels. We will construct the mammalian DEKA ion selectivity motif in NavAb, along with other potential determinants of ion selectivity, examine the significance of each amino acid substitution, and determine the 3D structure of the selectivity filter. (iii) We will reconstitute ast inactivation in NavAb and analyze its structural basis. The fast inactivation gate of mammalian Nav1.7 channels will be substituted as an intracellular linker in NavAb, and the amino acid residues forming the inactivation gate receptor will be introduced into the intracellular mouth of the pore. We will determine the functional properties and 3D structures of NavAb channels with a mammalian fast inactivation gate and inactivation gate receptor. (iv) Mutations that affect activation vs. inactivation of Nav1.7 channels cause the different hyperalgesic symptoms of erythromelalgia and paroxysmal extreme pain disorder. We will determine the structural basis for the differential effects of these mutations on Nav1.7 channels by introducing disease mutations into NavAb chimeras, confirming the functional impairments, and determining the 3D structures. These studies will provide essential structural information at the atomic level that wil elucidate mechanisms of voltage-dependent activation, ion conductance and selectivity, and fast inactivation of mammalian Nav channels. These studies will also provide a molecular template for understanding the pathophysiology of inherited channelopathies and for structure-based design of new generations of sodium channel therapeutics.

Public Health Relevance

Voltage-gated sodium channels initiate electrical activity in neurons and other excitable cells, and they are impaired in inherited epilepsy, migraine, chronic pain, and periodic paralysis. We will determine the three-dimensional structures of the key functional elements of sodium channels at the atomic level. This information will advance understanding of the molecular basis for neurological diseases, including chronic pain, and will provide a template for development of new generations of important drugs.

National Institute of Health (NIH)
Research Project (R01)
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Biophysics of Neural Systems Study Section (BPNS)
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Silberberg, Shai D
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University of Washington
Schools of Medicine
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Catterall, William A (2014) Sodium channels, inherited epilepsy, and antiepileptic drugs. Annu Rev Pharmacol Toxicol 54:317-38
Gamal El-Din, Tamer M; Scheuer, Todd; Catterall, William A (2014) Tracking S4 movement by gating pore currents in the bacterial sodium channel NaChBac. J Gen Physiol 144:147-57
Tang, Lin; Gamal El-Din, Tamer M; Payandeh, Jian et al. (2014) Structural basis for Ca2+ selectivity of a voltage-gated calcium channel. Nature 505:56-61
Catterall, William A (2014) Structure and function of voltage-gated sodium channels at atomic resolution. Exp Physiol 99:35-51
Gamal El-Din, Tamer M; Martinez, Gilbert Q; Payandeh, Jian et al. (2013) A gating charge interaction required for late slow inactivation of the bacterial sodium channel NavAb. J Gen Physiol 142:181-90
Chakrabarti, Nilmadhab; Ing, Christopher; Payandeh, Jian et al. (2013) Catalysis of Na+ permeation in the bacterial sodium channel Na(V)Ab. Proc Natl Acad Sci U S A 110:11331-6
Yarov-Yarovoy, Vladimir; DeCaen, Paul G; Westenbroek, Ruth E et al. (2012) Structural basis for gating charge movement in the voltage sensor of a sodium channel. Proc Natl Acad Sci U S A 109:E93-102
DeCaen, Paul G; Yarov-Yarovoy, Vladimir; Scheuer, Todd et al. (2011) Gating charge interactions with the S1 segment during activation of a Na+ channel voltage sensor. Proc Natl Acad Sci U S A 108:18825-30
Payandeh, Jian; Scheuer, Todd; Zheng, Ning et al. (2011) The crystal structure of a voltage-gated sodium channel. Nature 475:353-8
Catterall, William A (2010) Signaling complexes of voltage-gated sodium and calcium channels. Neurosci Lett 486:107-16

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