Linkage of the efflux-pump expression level with substrate extrusion rate in the MexAB-OprM efflux pump of Pseudomonas aeruginosa.
The amount of the subunit proteins of the MexAB-OprM efflux pump in Pseudomonas aeruginosa was quantified by the immunoblotting method. A single cell of the wild-type strain contained about 2500, 1000, and 1200 copies of MexA, MexB, and OprM, respectively, and their stoichiometry therefore was 2:1:1. The mexR mutant produced an eightfold higher level of these proteins than did wild-type cells. Assuming that MexB and OprM exist as a trimer in a pump assembly, the total number of MexAB-OprM per wild-type cell was calculated to be about 400 assemblies. The substrate efflux rate of MexAB-OprM was calculated from the fluorescent intensity of ethidium in intact cells that a single cell extruded ethidium at a maximum of about 3 x 10(-19) mol s(-1) and, therefore, the turnover rate of a single pump unit was predicted to be about 500 s(-1).
Narita
Shin-ichiro
S
Department of Molecular Life Science, Tokai University School of Medicine, Isehara 259-1193, Japan.
Eda
Shima
S
Yoshihara
Eisaku
E
Nakae
Taiji
T
D000900:Anti-Bacterial Agents / Q000494:pharmacology; D001425:Bacterial Outer Membrane Proteins / Q000378:metabolism*; D001692:Biological Transport; D002352:Carrier Proteins / Q000378:metabolism*; D002462:Cell Membrane / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D018432:Drug Resistance, Multiple; D004926:Escherichia coli / Q000378:metabolism; D004996:Ethidium / Q000494:pharmacology; D015151:Immunoblotting; D026901:Membrane Transport Proteins / Q000378:metabolism*; D009154:Mutation; D010957:Plasmids / Q000378:metabolism; D011550:Pseudomonas aeruginosa / Q000378:metabolism*; D013997:Time Factors; D015262:Xenobiotics / Q000494:pharmacology
D000900:Anti-Bacterial Agents; D001425:Bacterial Outer Membrane Proteins; D002352:Carrier Proteins; D026901:Membrane Transport Proteins; C118598:MexA protein, Pseudomonas aeruginosa; C118597:MexB protein, Pseudomonas aeruginosa; C402263:OprM protein, Pseudomonas aeruginosa; D015262:Xenobiotics; D004996:Ethidium