Crystal Structure of a Fibroblast Growth Factor Homologous Factor (FHF) Defines a Conserved Surface on FHFs for Binding and Modulation of Voltage-gated Sodium Channels

Regina Goetz,Katarzyna Dover,Fernanda Laezza,Nataly Shtraizent,Xiao Huang,Dafna Tchetchik,Anna V Eliseenkova,Chong-Feng Xu,Thomas A Neubert,David M Ornitz,Mitchell Goldfarb,Moosa Mohammadi

doi:10.1074/jbc.m109.001842

Abstract

Voltage-gated sodium channels (Nav) produce sodium currents that underlie the initiation and propagation of action potentials in nerve and muscle cells. Fibroblast growth factor homologous factors (FHFs) bind to the intracellular C-terminal region of the Nav alpha subunit to modulate fast inactivation of the channel. In this study we solved the crystal structure of a 149-residue-long fragment of human FHF2A which unveils the structural features of the homology core domain of all 10 human FHF isoforms. Through analysis of crystal packing contacts and site-directed mutagenesis experiments we identified a conserved surface on the FHF core domain that mediates channel binding in vitro and in vivo. Mutations at this channel binding surface impaired the ability of FHFs to co-localize with Navs at the axon initial segment of hippocampal neurons. The mutations also disabled FHF modulation of voltage-dependent fast inactivation of sodium channels in neuronal cells. Based on our data, we propose that FHFs constitute auxiliary subunits for Navs.

Highlights

Voltage-gated sodium channels (Nav)3 produce sodium currents that underlie the initiation and propagation of action potentials in nerve and muscle cells
For the cardiac channel Nav1.5, it has been proposed that interaction between the DIII-DIV loop and the C-terminal domain of the ␣ subunit stabilizes the inactivated state of the channel [5]
We present a crystal structure of FHF2A, which enabled us to define a conserved surface on FHFs required for binding to the C-terminal domain of Navs and for the modulation of channel inactivation

Summary

Introduction

Voltage-gated sodium channels (Nav)3 produce sodium currents that underlie the initiation and propagation of action potentials in nerve and muscle cells. We present a crystal structure of FHF2A, which enabled us to define a conserved surface on FHFs required for binding to the C-terminal domain of Navs and for the modulation of channel inactivation. To analyze FHF1 binding to the C-terminal domain of Nav ␣ subunit isoforms, FHF1A1–243 and FHF1B1–181 (Fig. 1A) were immobilized by amine coupling on two flow channels of a research grade CM5 chip (Biacore AB; ϳ91 fmol/mm2 of flow channel).

Results

Conclusion