KcsA potassium channel

The potassium crystallographically-sited activation channel (KcsA) is a prokaryotic potassium ion channel from the soil bacteria Streptomyces Lividans activated by changes in pH. Roderick MacKinnon and his colleagues were the first to crystallize a KcsA potassium channel.

KcsA has become the most extensively studied potassium channel, and is widely used as the template in potassium channel research.

KcsA Channel
KcsA is a tetramer composed of four identical subunits of two transmembrane helices (the outer-helix M1 and the inner-helix M2) linked by a reentrant loop, dispersed symmetrically around a common axis corresponding to the central pore. The pore can be divided into three parts: a selectivity filter near the extracellular side, a dilated water-filled cavity at the center, and a closed gate near the cytoplasmic side formed by four packed M2 helices. This architecture is found to be highly conserved in the potassium channel family, including both the eukaryotic and prokaryotic ones.

The KcsA channel is a proton-activated potassium channel that opens at acidic pH. It has two pH-sensing regions: 1) the charge cluster region at the boundary between the membrane and cytoplasm, and 2) the cytoplasmic domain. There is evidence to suggest that the main pH sensor is in the cytoplasmic domain. For example, Hirano et al showed that exchanging negatively charged amino acids for neutral ones made the KcsA channel insensitive to pH even though there were no amino-acid changes at the transmembrane region.

The KcsA channel is blocked by Cs+ ions and gating requires the presence of Mg2+ ions.

Structure
The structure of KcsA is that of a truncated cone, with a central pore running down the centre. The cone is made up of the M1 and M2 helices, which span the lipid bilayer. The wider end of the cone corresponds to the extracellular mouth of the channel. This envelops the pore (P) region, made up of the P-helices, plus a selectivity filter that is formed by a TVGYG sequence (Threonine, Valine, Glycine, Tyrosine, Glycine) characteristic of potassium channels. Beneath the selectivity filter is a central water-filled cavity. Finally, the pore-lining M2 helices constrict the intracellular mouth to form a putative gate region.

The KcsA channel is considered a model channel because the KcsA structure provides a framework for the understanding of Potassium selectivity and permeation and because it appears,from sequence comparisons, that the structure of this pore domain is conserved between diverse Potassium channels.

The X-ray structure of KcsA shows that there are two potassium ions within the selectivity filter (probably with a water molecule in between them), as well as a third Potassium ion in the central cavity. However, a crystal structure inevitably provides a static, spatially and temporally averaged image of a channel. To bridge the gap between molecular structure and physiological behavior, an understanding of the atomic resolution dynamics of potassium channels is required. One way in which this is approached is via simulation studies. Simulations reveal that interactions of potassium ions and water with the KcsA channel at both the selectivity filter and at the intracellular gate are dynamic. Potassium ions are unstable in the absence of other ions or molecules. In the cell cytoplasm, each potassium ion is surrounded by a shell of water. The cavity’s shape and size allows for approximately fifty molecules of water and one ion; X-ray crystallography has shown that water keeps potassium stable in the center of the cavity before the selectivity filter.

An aspect of KcsA that has not been fully addressed by simulation studies is that the crystal structure appears to be that of the ‘closed' form of the channel. This is reasonable as the closed state of the channel is favored at neutral pH, at which the crystal structure was solved. Electronic paramagnetic resonance (EPR) spectroscopic studies of KcsA suggest that channel opening is associated with changes in packing of the transmembrane helices, so as to widen the pore at its intracellular mouth.

The selectivity Filter
The X-ray structure of the KcsA Potassium channel revealed that selectivity filter is lined by backbone carbonyl groups from the residues of signature amino acid sequence TTVGYG, common to all known potassium channels. A dehydrated potassium ion fits in the narrow selectivity filter precisely so that the energetic costs and gains are well balanced. The main chain carbonyl oxygen atoms that make up the selectivity filter are held at a precise position that allows them to substitute for water molecules in the hydrated shell of the Potassium ion, but they are too far from the Sodium ion, which thereby retains its hydration reaction shell and is blocked from crossing through the pore. These observations led to a commonly accepted explanation of ion selectivity, which assumes that structural changes play the dominant role.

The selectivity filter of KcsA is formed by the backbone from four independent subunits usually labeled S1 to S4 starting at the extracellular side. The subunits are not bound directly to one another, and the result is a fairly flexible (liquid-like) pore structure.