The structure of the KtrAB potassium transporter

Abstract

In bacteria, archaea, fungi and plants the Trk, Ktr and HKT ion transporters are key components of osmotic regulation, pH homeostasis and resistance to drought and high salinity. These ion transporters are functionally diverse: they can function as Na + or K + channels and possibly as cation/K + symporters. They are closely related to potassium channels both at the level of the membrane protein and at the level of the cytosolic regulatory domains. Here we describe the crystal structure of a Ktr K + transporter, the KtrAB complex from Bacillus subtilis. The structure shows the dimeric membrane protein KtrB assembled with a cytosolic octameric KtrA ring bound to ATP, an activating ligand. A comparison between the structure of KtrAB-ATP and the structures of the isolated fulllength KtrA protein with ATP or ADP reveals a ligand-dependent conformational change in the octameric ring, raising new ideas about the mechanism of activation in these transporters.The Trk/Ktr/HKT superfamily of ion transporters includes the Trk, Ktr and HKT transporter families 1 . These transporters are closely related to the superfamily of tetrameric cation channels 2-4 , which includes potassium, sodium and calcium channels. The membrane proteins in both superfamilies share the same architecture 2,5-9 : four subunits or repeats, each with the TM-P loop-TM (where TM indicates transmembrane helix) structural motif first seen in the KcsA potassium channel structure 10 , assemble to form an ion pore with a 'selectivity filter' along its central axis. In addition, the cytosolic regulatory proteins of the Trk and Ktr transporters and of the MthK 11,12 and large-conductance Ca 2+ -gated (BK) K + channels [13][14][15] are RCK (regulate conductance of K+) domains.Ktr ion transporters are crucial K + transport systems in some bacteria 16,17 , with a role in resistance to osmotic stress and high salinity by mediating the early uptake of K + (refs 16, 18). The Ktr ion transporters are composed of two essential components 3,16 : a membrane protein (KtrB or KtrD) and a cytosolic regulatory protein (KtrA or KtrC). Studies with cells show that Ktr proteins 8,19 transport K + but are also permeable to Na + , and that K + transport is ATP 20 and Na + dependent 5,8,18 . Furthermore, it has been shown that truncated KtrA forms octameric rings 21,22 and that KtrB assembles as homodimers 6,21 . The structures described here clarify the molecular basis of some of these properties as well as the structural relationship between these transporters and ion channels.

A refined atomic scale model of the Saccharomyces cerevisiae K+-translocation protein Trk1p combined with experimental evidence confirms the role of selectivity filter glycines and other key residues

“…These residues were also the basis for matching pore helices and transmembrane helices in the earlier models. The correct alignment of the conserved selectivity filter glycine residues was confirmed by the crystal structures of TrkH and KtrB [14,15]. Mutagenesis studies with HKT and KtrB [40][41][42] showed that mutations of these glycines alter ion selectivity.…”

“…This helix is straight in K-channels. We wanted to exclude the possibility that this helix would also be a single straight helix in Trk1, therefore differing from the template structure [15]. The proline residues would -in case of a continued helix-represent a deviation from an optimal α-helix and could allow for function associated bending motions.…”

“…Recently, the crystal structures of Ktr/Trk/HKT-type transporters, TrkH [14] and KtrB [15], were solved. These structures supported earlier predictions about the similarity of their respective selectivity filters with that of K + channels and the conservation of the glycine residues [10].…”

Molecular evolutionary analysis of the high-affinity K+ transporter gene family in angiosperms

“…All of these sites were located in MPM domains (MPMa: 61V/T, 68S/G, 69M/L, 77F/L, and 82L/I; MPMb: 203F/S, 208T/S, 214F/L, and 236P/G; MPMc: 363Y/S; MPMd: 443F/L and 466G/S). Finally, functional analysis outcomes with TaHKT2;1, AtHKT1;1, and rice genes suggested that site 68S/G of the MPMa domain might play a central role in determining the Na + selectivity of the transporter (Horie et al, 2001;Vieira-Pires et al, 2013). After gene duplication, paralogs went through one of four processes: in the first (nonfunctionalization) one copy initially became a pseudogene and gradually became extinct (Petrov and Hartl, 2000).…”

“…We used the KtrAB potassium transporter from Bacillus subtilis (PDB id 4j7c) as our template (Vieira-Pires et al, 2013), which was obtained from the Protein Data Bank (http://www.pdb.org/pdb/home/home.do). Amino acid sequences were aligned with ClustalX v.1.83 (Thompson et al, 1997).…”