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Abstract:
The flow of ions across cell membranes facilitated by ion channels is an important function for all living cells. Despite the huge
amount of structural data provided by crystallography, elucidating the exact interactions between the selectivity filter atoms and bound ions is challenging. Here, we detect bound 15 N-labeled ammonium ions as a
mimic for potassium ions in ion channels using solid-state NMR under near-native conditions. The non-selective ion channel NaK showed two
ammonium peaks corresponding to its two ion binding sites, while its potassium-selective mutant NaK2K that has a signature potassium-
selective selectivity filter with four ion binding sites gave rise to four ammonium peaks. Ions bound in specific ion binding sites were identified based on magnetization transfer between the ions and carbon atoms in the selectivity filters. Magnetization transfer between bound ions and water molecules revealed that only one out of four ions in the selectivity filter of NaK2K is in close contact with water, which is in agreement with the direct knock-on ion conduction mechanism where ions are conducted through the channel by means of direct interactions without water molecules in between. Interestingly, the potassium-selective ion channels investigated here (NaK2K and, additionally, KcsA-Kv1.3) showed remarkably different chemical shifts for their bound ions, despite having identical amino acid sequences and crystal structures of their selectivity filters. Molecular dynamics simulations show similar ion binding and conduction behavior between ammonium and potassium ions and identify the origin of the differences between the investigated potassium channels
