Homology Modeling and Molecular Dynamics Simulation Studies of an Inward Rectifier Potassium Channel

Abstract

A homology model has been generated for the pore-forming domain of Kir6.2, a component of an ATP-sensitive K channel, based on the x-ray structure of the bacterial channel KcsA. Analysis of the lipid-exposed and pore-lining surfaces of the model reveals them to be compatible with the known features of membrane proteins and Kir channels, respectively. The Kir6.2 homology model was used as the starting point for nanosecond-duration molecular dynamics simulations in a solvated phospholipid bilayer. The overall drift from the model structure was comparable to that seen for KcsA in previous similar simulations. Preliminary analysis of the interactions of the Kir6.2 channel model with K(+) ions and water molecules during these simulations suggests that concerted single-file motion of K(+) ions and water through the selectivity filter occurs. This is similar to such motion observed in simulations of KcsA. This suggests that a single-filing mechanism is conserved between different K channel structures and may be robust to changes in simulation details. Comparison of Kir6.2 and KcsA suggests some degree of flexibility in the filter, thus complicating models of ion selectivity based upon a rigid filter.

Hetero-concatemeric KIR6.X4/SUR14 Channels Display Distinct Conductivities but Uniform ATP Inhibition

“…A similar idea, based on the hydrodynamic theory of Dwyer et al (19), has been used to explain a qualitatively similar effect of subunit order on g of heteromeric cyclic nucleotide-gated channels (20). With regard to possible effects on the dynamics of the selectivity filter, we note that the potential intersubunit salt bridges between the conserved R and E following the K ϩ channel signature G(Y/F)G sequence are unlikely as pK A calculations, based on a homology model of the K IR 6.2 tetramer embedded into a lipid bilayer (21), indicate that these E are protonated. Intersubunit H-bonds equivalent to those between Trp 68 in the pore helix and Tyr 78 in the signature motif of adjacent subunits in KcsA (22) are also unlikely because phenylalanines occupy the corresponding positions in K IR 6.X.…”

The ligand-sensitive gate of a potassium channel lies close to the selectivity filter

“…The equivalent residues in Kir6.2 are Thr 130 and Ile 131 ( Fig. 4A; Capener et al, 2000). Mutation of Thr 130 to valine in Kir6.2 did not result in functional channels, but mutation of Val 129 to threonine caused a marked decrease in Ba 2+ affinity: the IC 50 was 12 ± 0.2 µM (n = 5) compared with 0.8 ± 0.04 µM (n = 5) for the wild-type channel (Fig.…”

“…Support for this idea comes from the fact that the crystal structure of the bacterial K + channel MthK, imaged in the open state, reveals a widening of the pore at the helix-bundle crossing region (Jiang et al, 2002). Functional studies indicate that the activation gate of the voltage-dependent K + (K V ) channel Shaker is also located at the bundle crossing (Del Camino et al, 2000; The ligand-sensitive gate of a potassium channel lies close to the selectivity filter (A) Homology model of two of the four subunits of Kir6.2, generated by using the sequence alignment of Capener et al (2000) and the crystallographic coordinates of KcsA (Doyle et al, 1998). A Ba 2+ ion is shown to scale at the entrance to the selectivity filter.…”

Time-resolved Mechanism of Extracellular Gate Opening and Substrate Binding in a Glutamate Transporter

“…The van der Waals interactions were calculated using a cutoff distance of 10 Å, and long range electrostatic interactions were calculated using the particle mesh Ewald method. The single point charge model was adopted for water molecules as a plausible model for lipid simulations (21,22) along with the lipid parameters used in our previous MD simulations of other membrane proteins (23,24). The membrane consisted of 353 1-palmitoyl-2-oleoylphosphatidyl ethanolamine molecules, and the system was solvated with ϳ23,000 water molecules.…”