Restoring the function of hERG LQT2 N-terminal mutants with non-canonical amino acids.

Abstract

The human Ether-a-go-go related (hERG) potassium channels produce the IKr cardiac current, which drives the repolarization of the membrane potential and terminates action potentials in the heart. Point mutations that introduce a TAG stop codon in hERG channels have been found in patients with Long QT syndrome Type 2. LQTS2 patients have an increased risk of cardiac arrhythmias that may result in sudden cardiac arrest. The prolonged action potential duration (APD) that characterizes LQT2 can be explained by the loss of hERG function and thus a decreased IKr. This work studies the effect of TAG mutations on the expression and electrical properties of hERG channels and the rescue of function by incorporating non-canonical amino acids (ncAAs) in hERG TAG LQT2 mutants. When transfected in HEK293 cells, hERG1a-Q81TAG produces a functional channel with a faster deactivation rate and smaller currents than the wild type (WT) channel, while hERG1a-S182TAG, E229TAG and W398TAG transfected cells do not produce measurable currents. Using an ortholog set of aaRS and tRNA, we incorporated the ncAA L-ANAP into hERG TAG LQT2 mutants. We found that the incorporation of L-ANAP into hERG1a-Q81TAG restores the WT phenotype of hERG currents. Meanwhile, the incorporation of a ncAA into hERG1a-S182TAG and E229TAG, but not in W398TAG, results in the production of a full-length WT-like channel. We propose that the effect of hERG1a TAG mutants in LQTS2 is due to changes in channel kinetics or a diminished channel expression and that the WT channel function and expression can be rescued by the incorporation of a ncAA. This method may be useful in rescuing other channels or proteins with disease-linked TAG mutations.