Functional characterization of Actinobacillus pleuropneumoniae urea transport protein, ApUT.
Urea transporters (UTs) effect rapid flux of urea across biological membranes. In the mammalian kidney, UT activity is essential for effective urine concentration. In bacteria, UT-mediated urea uptake permits intracellular urease to degrade urea to ammonia and CO(2), a process that either buffers acid loads or provides nutrient nitrogen. We have characterized the urea transport channel protein ApUT from Actinobacillus pleuropneumoniae. Kinetic analysis of bacterial inside-out membranes enriched in ApUT showed approximately 28-fold increase in urea permeability (3.3 +/- 0.4 x 10(-4) cm/s) compared with control vesicles (0.11 +/- 0.02 x 10(-4) cm/s). In addition to urea, ApUT also conducts water. Urea and water transport across the channel was phloretin and mercury inhibitable, and the site of inhibition may be located on the cytoplasmic side of the protein. Glycerol and urea analogs, such as methylamine, dimethylurea, formamide, acetamide, methylurea, propanamide, and ethylamine did not permeate across ApUT.
Godara
Geeta
G
Beth Israel Deaconess Medical Center, Cambridge, MA 02139, USA.
Smith
Craig
C
Bosse
Janine
J
Zeidel
Mark
M
Mathai
John
J
D016977:Actinobacillus pleuropneumoniae / Q000235:genetics / Q000378:metabolism*; D001426:Bacterial Proteins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D001692:Biological Transport; D002463:Cell Membrane Permeability; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000378:metabolism; D005990:Glycerol / Q000378:metabolism; D007700:Kinetics; D026901:Membrane Transport Proteins / Q000187:drug effects / Q000235:genetics / Q000378:metabolism*; D008627:Mercuric Chloride / Q000494:pharmacology; D010693:Phloretin / Q000494:pharmacology; D014508:Urea / Q000031:analogs & derivatives / Q000378:metabolism*; D014867:Water / Q000378:metabolism
D001426:Bacterial Proteins; D026901:Membrane Transport Proteins; D014867:Water; D008627:Mercuric Chloride; D014508:Urea; D005990:Glycerol; D010693:Phloretin