A hydrophobic nexus at the heart of hERG K channel gating

Main text

The human ether á go-go related gene (hERG) is a voltage-activated potassium channel of critical importance in the human heart, where it drives repolarization of cardiac action potentials (1,2,3,4). hERG opening and closing (gating) is tuned for its role in heart, and molecular specializations in hERG include an intracellular N-terminal PAS (Per-Arnt-Sim) domain (5) and a C-terminal cyclic nucleotide binding homology domain (CNBHD) (6) (Fig. 1). In hERG, the PAS domain regulates several biophysical properties including inactivation, steady-state activation, and, most robustly, channel closing (deactivation), as deletion or disruption of the PAS by point mutations or disease-causing mutations speeds up deactivation by as much as fivefold (1,7,8,9,10,11,12,13,14,15,16). The PAS domain makes a direct interaction with the CNBHD from an adjacent subunit and this interaction is required to regulate gating (8,10) (Fig. 1). Isolated structures of the PAS indicated that it can be divided into subdomains including the globular PAS, PAS-Cap alpha helix, and PAS-Cap tip, and mutations in each subdomain also disrupted gating (12,17,18,19,20) (Fig. 1). In the context of all four channel subunits, the PAS-Cap tip is positioned to point toward the transmembrane domain gating machinery, indicating a possible regulatory mechanism (21) (Fig. 1). But, despite intense scrutiny, many aspects of the mechanism for PAS domain regulation of hERG gating are unclear.

Figure 1.

Schematic of hydrophobic nexus and repositioning of the PAS-Cap tip by mutations in the PAS-Cap helix. Two (of four) hERG subunits showing the S1–S6 transmembrane domains and the location of the hydrophobic nexus formed between the intracellular globular PAS (cyan), PAS-Cap helix, and CNBHD. The PAS-Cap tip and amino acid V3 are positioned nearby the S4-S5 linker (green) and amino acid D540. (Inset) Schematic of V3 relative to D540 and distance change in V3 (denoted by V3^∗ and red arrow) in the repositioned PAS-Cap tip (dashed orange) due to point mutations at residues in the hydrophobic nexus (I19S, I18S, and L15S) in the PAS-Cap helix.

New insights come from a recent article in Biophysical Journal (22), in which the Robertson lab makes several key discoveries about the PAS domain regulatory mechanism. First, they identify a grouping of hydrophobic amino acids from different domains including the globular PAS (cyan), CNBHD (pink), and PAS-Cap helix (orange), which they term the “hydrophobic nexus” (Fig. 1). Second, they show that point mutations at selected hydrophobic residues within the hydrophobic nexus disrupt the mechanism, as determined by accelerated deactivation kinetics measured with electrophysiology (22). hERG channel deactivation is characteristically very slow, for instance deactivation takes approximately 500 ms at −120 mV, but is accelerated up to threefold by point mutations in the hydrophobic nexus (22). The hydrophobic nexus is thus a new necessary component in the mechanism of hERG channel deactivation.

But how do the hydrophobic nexus mutations perturb the deactivation mechanism? To address this, the authors used molecular dynamics simulations. They show that point mutations in the PAS-Cap helix change the position of the adjacent PAS-Cap tip relative to the S4-S5 linker (Fig. 1, inset). In particular, the distance from amino acid V3 in the PAS-Cap tip to amino acid D540 in the S4-S5 linker was increased by point mutations L15S, I18S, or I19S in the PAS-Cap helix. The implication is that the PAS-Cap tip was repositioned, relative to the S4-S5 linker, by a distance of a few angstroms (Fig. 1, inset, dashed lines), and they proposed that the increase in the distance of the PAS-Cap tip from the transmembrane gating machinery was responsible for the acceleration of channel deactivation gating. The broader implications of the study are that while point mutations in one region (e.g., the PAS-Cap helix) certainly alter the chemistry of the local side chain, they can also affect the three-dimensional positioning of adjacent regions (e.g., the PAS-Cap tip). This is a consideration for other studies that use point mutations and the interpretation of results.

These findings add another biophysical piece to the puzzle of hERG deactivation gating. Remarkably, the results can be inferred to mean that angstrom-scale motions of an extended peptide (the PAS-Cap tip) are vital for hERG gating and, by extension, maintenance of cardiac repolarization properties that are critical for normal human cardiac function.

Editor: Marcel Goldschen-Ohm.