PDBsum entry 2atk

Immune system/ion transport

PDB id: 2atk

Contents

Protein chains

219 a.a. *

212 a.a. *

103 a.a. *

Ligands

F09

Metals

__K ×7

Waters ×43

* Residue conservation analysis

PDB id:

2atk

Name:

Immune system/ion transport

Title:

Structure of a mutant kcsa k+ channel

Structure:

Antibody fab fragment heavy chain. Chain: a. Engineered: yes. Antibody fab fragment light chain. Chain: b. Engineered: yes. Voltage-gated potassium channel. Chain: c. Synonym: potassium channel kcsa.

Source:

Mus musculus. House mouse. Organism_taxid: 10090. Streptomyces lividans. Organism_taxid: 1916. Expressed in: escherichia coli. Expression_system_taxid: 562

Biol. unit:

Dodecamer (from PQS)

Resolution:

2.50Å R-factor: 0.238 R-free: 0.261

Authors:

J.F.Cordero-Morales,L.G.Cuello,Y.Zhao,V.Jogini,S.Chakrapani,B.Roux, E.Perozo

Key ref:

J.F.Cordero-Morales et al. (2006). Molecular determinants of gating at the potassium-channel selectivity filter. Nat Struct Mol Biol, 13, 311-318. PubMed id: 16532009 DOI: 10.1038/nsmb1069

Date:

25-Aug-05 Release date: 07-Mar-06

Protein chain

No UniProt id for this chain

Struc: 219 a.a.

Protein chain

No UniProt id for this chain

Struc: 212 a.a.

Protein chain

P0A334  (KCSA_STRLI) -  pH-gated potassium channel KcsA from Streptomyces lividans

Seq: 160 a.a.

Struc: 103 a.a.*

* PDB and UniProt seqs differ at 2 residue positions (black crosses)

DOI no: 10.1038/nsmb1069

Nat Struct Mol Biol 13:311-318 (2006)

PubMed id: 16532009

Molecular determinants of gating at the potassium-channel selectivity filter.

J.F.Cordero-Morales, L.G.Cuello, Y.Zhao, V.Jogini, D.M.Cortes, B.Roux, E.Perozo.

ABSTRACT

We show that in the potassium channel KcsA, proton-dependent activation is followed by an inactivation process similar to C-type inactivation, and this process is suppressed by an E71A mutation in the pore helix. EPR spectroscopy demonstrates that the inner gate opens maximally at low pH regardless of the magnitude of the single-channel-open probability, implying that stationary gating originates mostly from rearrangements at the selectivity filter. Two E71A crystal structures obtained at 2.5 A reveal large structural excursions of the selectivity filter during ion conduction and provide a glimpse of the range of conformations available to this region of the channel during gating. These data establish a mechanistic basis for the role of the selectivity filter during channel activation and inactivation.

Selected figure(s)

Figure 4.
Figure 4. Two crystal structures of the E71A mutant. (a) Electron density map of residues 60–84 from two diagonally symmetric subunits for the crystal form of E71A with residue Asp80 flipped upward (flipped E71A). Sticks, polypeptide chain; blue mesh, 1- contour of the 2F[o] – F[c] electron density map for the protein; magenta mesh, 2- contour of the ions; black dotted ovals, cavities created by the absence of the Glu71 side chain. (b) Single-subunit line representation of the P-loop of the flipped (red) and nonflipped (black) E71A structures overlaid onto the wild-type structure^32 (PDB entry 1K4C; gray) highlights the conformational rearrangements in the backbone of the selectivity filter (residues 75–79). Insets, the side chain conformational changes in Asp80 and Trp67 as fitted to the 2- contour of the simulated annealing omit map. The omit maps were calculated for residues 79–84 and 67, respectively; atoms within 3.5 Å of selected residues were also omitted in the calculation. The density attributed to the alternate rotamer of the Trp67 side chain in the flipped X-ray structure is clearly distinct from the density of the lipid observed near this position in the WT X-ray structure^33. (c) One-dimensional electron density profiles for the two crystal conformations of the E71A mutant (flipped and nonflipped). Top, F[o] – F[c] omit maps of K^+ ions in the selectivity filter shown relative to the protein model. The electron density maps are shown as a 6- contour for the flipped E71A conformer and as 7- (blue) and 4- (cyan) contours for the nonflipped E71A conformer. Different contour levels were chosen for the purpose of visual clarity. Bottom, one-dimensional electron density profile along the central symmetry (z) axis is shown using the ion in the cavity as z = 0. Gray-shaded peaks represent the profile for the wild-type channel (PDB entry 1K4C at 2.0 Å). Numbers and E at top denote the K^+-binding sites (S0–S4 and S[ext], respectively).(d) Comparison of crystallographic B-factors for P-loop residues (63–83) from the WT KcsA structure (bottom chart) and the nonflipped E71A mutant (E71A-NF, top chart). Black dotted line represents the mean value for all atoms in the P-loop. Vertical capped lines represent the 3 values for each data set.

Figure 6.
Figure 6. A mechanistic interpretation of KcsA gating. (a) Possible mechanism of action of the E71A mutation in stabilizing the open state. A single-subunit P-loop is shown with positions 67, 71 and 80 in stick representation. In the wild-type channel (left), the interaction between Asp80 and Trp67 destabilizes the conductive conformation of the filter and promotes inactivation through an as yet unknown mechanism. Eliminating the Asp80-Glu71 carboxyl-carboxylate (E71A, right) disrupts the hydrogen bonding network between the signature sequence (Gly-Tyr-Gly-Asp) and the pore helix, causing an increase in Asp80 dynamics and perturbing the Asp80-Trp67 interaction. This sharply decreases entry into the inactivated state, stabilizing the open state. (b) Top, cartoon representation of the structural conformation associated with each kinetic state. Bottom, correlation of specific kinetic transitions with KcsA single-channel behavior. Because stationary gating is dominated by the deeply inactivated state, single-channel openings occur mainly as a result of rare returns from the inactivated state owing to conformational changes in the selectivity filter while the lower gate remains open.

The above figures are reprinted by permission from Macmillan Publishers Ltd: Nat Struct Mol Biol (2006, 13, 311-318) copyright 2006.

Figures were selected by an automated process.