The projection structure of Kch, a putative potassium channel in Escherichia coli, by electron crystallography.
The kch gene, the only potassium channel gene in Escherichia coli, has the property to express both full-length Kch and its cytosolic domain (RCK) due to a methionine at position 240. The RCK domains are believed to form an octameric ring structure and regulate the gating of the potassium channels after having bound certain ligands. Several different gating ring structures have been reported for the soluble RCK domains, however, these were studied isolated from their transmembrane parts. We previously reported an octameric structure of Kch in solution by electron microscopy and single particle reconstruction, composed of two tetrameric full-length proteins through RCK interaction. To exclude the effect of the detergent, we have now performed an electron crystallographic study of the full-length Kch in membrane bound form. Well-ordered two-dimensional crystals were grown in a natural phospholipid environment. A projection map merged from the fifteen best images extended to 6Å resolution. The c12 two-sided plane group of the two-dimensional crystals showed that Kch crystallized as two symmetrically related overlapping layers. The arrangement suggests that the two layers of RCK domains are shifted with respect to each other and the RCK octameric gating ring of Kch does not form under the crystallization condition.
Karolinska Institutet, Dept of Biosciences and Nutrition and KTH Royal Institute of Technology, School of Technology and Health, Novum, S-14183 Huddinge, Sweden. Electronic address: Qie.Kuang@ki.se.
Qie
Q
Kuang
Pasi
P
Purhonen
Caroline
C
Jegerschöld
Hans
H
Hebert
D000595:Amino Acid Sequence; D003461:Crystallography / Q000379:methods; D004583:Electrons; D004926:Escherichia coli / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D029968:Escherichia coli Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D015640:Ion Channel Gating; D008958:Models, Molecular; D008969:Molecular Sequence Data; D015221:Potassium Channels / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D055503:Protein Multimerization; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism
D029968:Escherichia coli Proteins; D015221:Potassium Channels; D011994:Recombinant Proteins; C101871:kch protein, E coli
Electron microscopy; Membrane protein; Potassium channel; Projection map; RCK