The cell wall of the pathogenic bacterium Rhodococcus equi contains two channel-forming proteins with different properties.
We have identified in organic solvent extracts of whole cells of the gram-positive pathogen Rhodococcus equi two channel-forming proteins with different and complementary properties. The isolated proteins were able to increase the specific conductance of artificial lipid bilayer membranes made from phosphatidylcholine-phosphatidylserine mixtures by the formation of channels able to be permeated by ions. The channel-forming protein PorA(Req) (R. equi pore A) is characterized by the formation of cation-selective channels, which are voltage gated. PorA(Req) has a single-channel conductance of 4 nS in 1 M KCl and shows high permeability for positively charged solutes because of the presence of negative point charges. According to the results of sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), the protein has an apparent molecular mass of about 67 kDa. The analysis (using the effect of negative charges on channel conductance) of the concentration dependence of the single-channel conductance suggested that the diameter of the cell wall channel is about 2.0 nm. The second channel (formed by PorB(Req) [R. equi pore B]) shows a preferred movement of anions through the channel and is not voltage gated. This channel shows a single-channel conductance of 300 pS in 1 M KCl and is characterized by the presence of positive point charges in or near the channel mouth. Based on SDS-PAGE, the apparent molecular mass of the channel-forming protein is about 11 kDa. Channel-forming properties of the investigated cell wall porins were compared with those of others isolated from mycolic acid-containing actinomycetes. We present here the first report of a fully characterized anion-selective cell wall channel from a member of the order Actinomycetales.
Riess
Franziska G
FG
Lehrstuhl für Biotechnologie, Biozentrum der Universität Würzburg, Am Hubland, D-97074 Würzburg, Germany.
Elflein
Marion
M
Benk
Michael
M
Schiffler
Bettina
B
Benz
Roland
R
Garton
Natalie
N
Sutcliffe
Iain
I
D000818:Animals; D001426:Bacterial Proteins; D015153:Blotting, Western; D002473:Cell Wall / Q000737:chemistry*; D006801:Humans; D007473:Ion Channels / Q000032:analysis*; D008051:Lipid Bilayers / Q000737:chemistry; D008564:Membrane Potentials; D008970:Molecular Weight; D010713:Phosphatidylcholines; D010718:Phosphatidylserines; D018272:Porins / Q000737:chemistry / Q000302:isolation & purification*; D016929:Rhodococcus equi / Q000737:chemistry*
D001426:Bacterial Proteins; D007473:Ion Channels; D008051:Lipid Bilayers; D010713:Phosphatidylcholines; D010718:Phosphatidylserines; C419549:PorB protein, Chlamydia trachomatis; D018272:Porins; C057699:porin protein, Neisseria