Analysis of a putative voltage-gated prokaryotic potassium channel.
Most of the completely sequenced prokaryotic genomes contain genes of potassium channel homologues, but there is still not much known about the role of these proteins in prokaryotes. Here we describe the large-scale overproduction and purification of a prokaryotic voltage-gated potassium channel homologue, Kch, from Escherichia coli. After successful overproduction of the protein, a specific increase in the potassium permeability of the cells was found. Kch could be purified in large amounts using classical purification methods to prevent aggregation of the protein. The physiological state of the protein was revealed to be a homotetramer and the protein was shown to be localized to the cytoplasmic membrane of the cells. In the course of the localization studies, we found a specific increase in the density of the cytoplasmic membrane on Kch production. This was linked to the observed increase in the protein to lipid ratio in the membranes. Another observed change in the membrane composition was an increase in the cardiolipin to phosphatidylglycerol ratio, which may indicate a specific cardiolipin requirement of Kch. On the basis of some of our results, we discuss a function for Kch in the maintenance of the membrane potential in E. coli.
Ungar
D
D
Department of Molecular Membrane Biology, Max-Planck-Institute of Biophysics, Frankfurt/Main, Germany.
Barth
A
A
Haase
W
W
Kaunzinger
A
A
Lewitzki
E
E
Ruiz
T
T
Reiländer
H
H
Michel
H
H
D002462:Cell Membrane / Q000737:chemistry / Q000648:ultrastructure; D002942:Circular Dichroism; D004926:Escherichia coli / Q000737:chemistry / Q000378:metabolism; D029968:Escherichia coli Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D015640:Ion Channel Gating; D011188:Potassium / Q000378:metabolism; D015221:Potassium Channels / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D020836:Protein Structure, Quaternary; D017433:Protein Structure, Secondary; D011994:Recombinant Proteins; D012995:Solubility; D017550:Spectroscopy, Fourier Transform Infrared
D029968:Escherichia coli Proteins; D015221:Potassium Channels; D011994:Recombinant Proteins; C101871:kch protein, E coli; D011188:Potassium