PDBsum entry 2k1e

Membrane protein

PDB id: 2k1e

Contents

Protein chains

103 a.a. *

* Residue conservation analysis

PDB id:

2k1e

Name:

Membrane protein

Title:

Nmr studies of a channel protein without membranes: structure and dynamics of water-solubilized kcsa

Structure:

Water soluble analogue of potassium channel, kcsa. Chain: a, b, c, d. Engineered: yes

Source:

Escherichia coli. Expressed in: escherichia coli. Other_details: designed variant of kcsa

NMR struc:

20 models

Authors:

D.Ma,Y.Xu,T.Tillman,P.Tang,E.Meirovitch,R.Eckenhoff,A.Carnini

Key ref:

D.Ma et al. (2008). NMR studies of a channel protein without membranes: structure and dynamics of water-solubilized KcsA. Proc Natl Acad Sci U S A, 105, 16537-16542. PubMed id: 18948596 DOI: 10.1073/pnas.0805501105

Date:

29-Feb-08 Release date: 11-Nov-08

Protein chains

No UniProt id for this chain

Struc: 103 a.a.

DOI no: 10.1073/pnas.0805501105

Proc Natl Acad Sci U S A 105:16537-16542 (2008)

PubMed id: 18948596

NMR studies of a channel protein without membranes: structure and dynamics of water-solubilized KcsA.

D.Ma, T.S.Tillman, P.Tang, E.Meirovitch, R.Eckenhoff, A.Carnini, Y.Xu.

ABSTRACT

Structural studies of polytopic membrane proteins are often hampered by the vagaries of these proteins in membrane mimetic environments and by the difficulties in handling them with conventional techniques. Designing and creating water-soluble analogues with preserved native structures offer an attractive alternative. We report here solution NMR studies of WSK3, a water-soluble analogue of the potassium channel KcsA. The WSK3 NMR structure (PDB ID code 2K1E) resembles the KcsA crystal structures, validating the approach. By more stringent comparison criteria, however, the introduction of several charged residues aimed at improving water solubility seems to have led to the possible formations of a few salt bridges and hydrogen bonds not present in the native structure, resulting in slight differences in the structure of WSK3 relative to KcsA. NMR dynamics measurements show that WSK3 is highly flexible in the absence of a lipid environment. Reduced spectral density mapping and model-free analyses reveal dynamic characteristics consistent with an isotropically tumbling tetramer experiencing slow (nanosecond) motions with unusually low local ordering. An altered hydrogen-bond network near the selectivity filter and the pore helix, and the intrinsically dynamic nature of the selectivity filter, support the notion that this region is crucial for slow inactivation. Our results have implications not only for the design of water-soluble analogues of membrane proteins but also for our understanding of the basic determinants of intrinsic protein structure and dynamics.

Selected figure(s)

Figure 1.
Comparison of WSK3 with KcsA. (A and B) The averaged NMR structure (green in A) and the 20 lowest energy structures (black in B) of WSK3 are superimposed on the low-K^+ KcsA crystal structure (1K4D in light gray). In A, the mutations made to facilitate water solubility and agitoxin-2 binding are highlighted in orange and black, respectively. (C) Sequence alignment and relative numbering of KcsA and WSK3. The mutations are highlighted in gold. The selectivity filter is enclosed in the red rectangle. The kink near V85 in WSK3 is marked with an asterisk. The underlined residues are non-α-helix in some structures.

Figure 3.
Stabilization of the selectivity filter and tetramer conformation in WSK3 by a network of salt bridges and hydrogen bonds. Important side chains are depicted in the licorice representation, with hydrogen bonds indicated by red dashed lines. (A and B) The same region is depicted from different viewing angles. (C) Quaternary relationship among W46, W47, E50, Y57, and R68 in WSK3. W67 and W68 in KcsA are shown in black lines for comparison. (D) Comparison of the selectivity filter between WSK3 structure and KcsA crystal structures obtained in the presence of high (1K4C) and low (1K4D) K^+ concentrations. WSK3 is depicted in thick sticks and KcsA in thin lines. Element colors: C, gray; N, blue; O, red; and K, yellow. K^+ locations are taken from the crystal structures and not from the NMR data.

Figures were selected by an automated process.