Molecular Biophysics
Structural properties determining low K+ affinity of the selectivity filter in the TWIK1 K+ channel

https://doi.org/10.1074/jbc.RA118.001817Get rights and content
Under a Creative Commons license
open access

Canonical K+ channels are tetrameric and highly K+-selective, whereas two-pore–domain K+ (K2P) channels form dimers, but with a similar pore architecture. A two-pore–domain potassium channel TWIK1 (KCNK1 or K2P1) allows permeation of Na+ and other monovalent ions, resulting mainly from the presence of Thr-118 in the P1 domain. However, the mechanistic basis for this reduced selectivity is unclear. Using ion-exchange–induced difference IR spectroscopy, we analyzed WT TWIK1 and T118I (highly K+-selective) and L228F (substitution in the P2 domain) TWIK1 variants and found that in the presence of K+ ions, WT and both variants exhibit an amide-I band at 1680 cm−1. This band corresponds to interactions of the backbone carbonyls in the selectivity filter with K+, a feature very similar to that of the canonical K+ channel KcsA. Computational analysis indicated that the relatively high frequency for the amide-I band is well explained by impairment of hydrogen bond formation with water molecules. Moreover, concentration-dependent spectral changes indicated that the K+ affinity of the WT selectivity filter was much lower than those of the variants. Furthermore, only the variants displayed a higher frequency shift of the 1680-cm−1 band upon changes from K+ to Rb+ or Cs+ conditions. High-speed atomic force microscopy disclosed that TWIK1's surface morphology largely does not change in K+ and Na+ solutions. Our results reveal the local conformational changes of the TWIK1 selectivity filter and suggest that the amide-I bands may be useful “molecular fingerprints” for assessing the properties of other K+ channels.

potassium channel
metal ion-protein interaction
infrared spectroscopy (IR spectroscopy)
biophysics
Fourier transform IR (FTIR)
ion channel
molecular dynamics
quantum chemistry
fluorescence

Cited by (0)

This work was supported in part by the Japan Society for the Promotion of Science KAKENHI Grants 25840122 and 17K15109 (to H. T.); 16H00778, 16KT0165, and 17K05757 (to M. H.); and 22247024, 22770159, 24650203, 26640047, and 26708002 (to Y. F.); NINS program for cross-disciplinary study (to Y. F. and K. N.); the Cooperative Study Program of National Institute for Physiological Sciences; and JST CREST Grant JPMJCR17N5. The authors declare that they have no conflicts of interest with the contents of this article.

This article contains Figs. S1–S7, supporting Methods, supporting Movies S1–S11, and supporting Refs. 1–2.

1

Present address: Dept. of Physiology, Division of Life Sciences, Faculty of Medicine, Osaka Medical College, 2-7 Daigaku-machi, Takatsuki, Osaka 569-8686, Japan.