Structures of the T cell potassium channel Kv1.3 with immunoglobulin modulators

Abstract

The Kv1.3 potassium channel is expressed abundantly on activated T cells and mediates the cellular immune response. This role has made the channel a target for therapeutic immunomodulation to block its activity and suppress T cell activation. We determined structures of human Kv1.3 alone, with a nanobody inhibitor, and with an antibody-toxin fusion blocker. Rather than block the channel directly, four copies of the nanobody bind the tetramer’s voltage sensing domains and the pore domain to induce an inactive pore conformation. In contrast, the antibody-toxin fusion docks its toxin domain at the extracellular mouth of the channel to insert a critical lysine into the pore. The lysine induces an active conformation of the pore yet blocks ion permeation. This study visualizes Kv1.3 pore dynamics, defines two distinct mechanisms to suppress Kv1.3 channel activity with exogenous inhibitors, and provides a framework to aid development of emerging T cell immunotherapies.

Driving Forces underlying Selectivity Filter Gating in the MthK Potassium Channel

“…Similar asymmetric SF conformations to ones reported in this work have been also observed in the structures of some two-pore domain (K2P) potassium channels: TREK-1 [40] and TASK-2 [41]. Specifically, we highlight that the conformational transitions (inactivation) observed in our MD simulations of both MthK WT and V55E channels involve widening (dilation) of the SF at the level of ‘second’ glycines (G63 in MthK), in agreement with available structural data for Shaker W434F, Kv1.2 W362F and WT Kv1.3 channels [27,29,30].…”

“…Structures of KcsA, stably-inactivated mutants of voltage-gated K + channels ( Shaker W434F and Kv1.2 W362F) and WT Kv1.3 channels revealed that C-type inactivation-related conformational changes at the SF can follow at least two distinct paths ( Fig. 6 ) [27–30]: in KcsA the SF pinches (constricts) at the level of the ‘first’ glycine (G77, corresponding to G61 in MthK), which is accompanied by flipping of valine (V76 in KcsA, corresponding to V60 in MthK) carbonyl away from the SF axis. The SF also loses two K + ions during inactivation, and has remaining ions bound at S1 and S4, whereas S3 is occupied by a water molecule, which presumably enhances the valine flipping.…”

“…Our previous work on the Kv1.2 channel had predicted aspartate flipping, SF dilation and loss of K + ions from S0-S2, also using the CHARMM force field [31], before recent cryoEM structures of stably-inactivated Shaker W434F, Kv1.2 W362F and Kv1.3 channels became available ( Fig. 6 C-F ) [27,29,30]. In MthK, previous MD simulations done with the CHARMM force field (and reduced interaction strength between K + ions and carbonyl oxygens) revealed that K + ions from sites S1, S2, and S3 need to unbind before the SF collapse, which was then structurally similar to the one seen in KcsA, having a constriction at the level of the ‘first’ glycine (G61), V60 and G61 carbonyl flipping and increased number of water molecules behind the SF [32].…”

Mechanism of 4-aminopyridine inhibition of the lysosomal channel TMEM175

“…Cryo-EM maps and atomic coordinates for 4-AP bound TMEM175 have been deposited with the European Molecular Data Bank (EMDB, https://www.ebi.ac.uk/emdb/ ) and Protein Data Bank (PDB, https://www.rcsb.org/ ) (accession nos. EMDB-27436 ( 42 ) and PDB 8DHM ( 43 )) Datasets from the PDB used in this study include 2A79 ( 44 ), 6WC9 ( 45 ), 7PHL ( 46 ), 7SSV ( 47 ), and 7UNL ( 48 ). Force field parameters for 4-AP compatible with CHARMM are publicly available at https://github.com/Faraldo-Gomez-Lab-at-NIH/Download .…”