NMR Structural Analysis of Isolated Shaker Voltage-Sensing Domain in LPPG Micelles

Hongbo Chen, Junkun Pan, Disha M. Gandhi, C. Dockendorff, Q. Cui, B. Chanda, Katherine A. Henzler-Wildman

Research output: Contribution to journalArticlepeer-review

2 Scopus citations

Abstract

The voltage-sensing domain (VSD) is a conserved structural module that regulates the gating of voltage-dependent ion channels in response to a change in membrane potential. Although the structures of many VSD-containing ion channels are now available, our understanding of the structural dynamics associated with gating transitions remains limited. To probe dynamics with site-specific resolution, we utilized NMR spectroscopy to characterize the VSD derived from Shaker potassium channel in 1-palmitoyl-2-hydroxy-sn-glycero-3-phospho-(1'-rac-glycerol) (LPPG) micelles. The backbone dihedral angles predicted based on secondary chemical shifts using torsion angle likeliness obtained from shift (TALOS+) showed that the Shaker-VSD shares many structural features with the homologous Kv1.2/2.1 chimera, including a transition from α-helix to 310 helix in the C-terminal portion of the fourth transmembrane helix. Nevertheless, there are clear differences between the Shaker-VSD and Kv1.2/2.1 chimera in the S2-S3 linker and S3 transmembrane region, where the organization of secondary structure elements in Shaker-VSD appears to more closely resemble the KvAP-VSD. Comparison of microsecond-long molecular dynamics simulations of Kv 1.2-VSD in LPPG micelles and a 1-palmitoyl-2-oleoyl-glycero-3-phosphocholine (POPC) bilayer showed that LPPG micelles do not induce significant structural distortion in the isolated voltage sensor. To assess the integrity of the tertiary fold, we directly probed the binding of BrMT analog 2-[2-({[3-(2-amino-ethyl)-6-bromo-1H-indol-2-yl]methoxy}k7methyl)-6-bromo-1H-indol-3-yl]ethan-1-amine (BrET), a gating modifier toxin, and identified the location of the putative binding site. Our results suggest that the Shaker-VSD in LPPG micelles is in a native-like fold and is likely to provide valuable insights into the dynamics of voltage-gating and its regulation.

Original languageEnglish
Pages (from-to)388-398
Number of pages11
JournalBiophysical Journal
Volume117
Issue number2
DOIs
StatePublished - Jul 23 2019

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