Purification and activation properties of UreD-UreF-urease apoprotein complexes.
In vivo assembly of the Klebsiella aerogenes urease nickel metallocenter requires the presence of UreD, UreF, and UreG accessory proteins and is further facilitated by UreE. Prior studies had shown that urease apoprotein exists in an uncomplexed form as well as in a series of UreD-urease (I.-S. Park, M.B. Carr, and R.P. Hausinger, Proc. Natl. Acad. Sci. USA 91:3233-3237, 1994) and UreD-UreF-UreG-urease (I.-S. Park and R.P. Hausinger, J. Bacteriol. 177:1947-1951, 1995) apoprotein complexes. This study demonstrates the existence of a distinct series of complexes consisting of UreD, UreF, and urease apoprotein. These novel complexes exhibited activation properties that were distinct from urease and UreD-urease apoprotein complexes. Unlike the previously described species, the UreD-UreF-urease apoprotein complexes were resistant to inactivation by NiCl2. The bicarbonate concentration dependence for UreD-UreF-urease apoenzyme activation was significantly decreased compared with that of the urease and UreD-urease apoproteins. Western blot (immunoblot) analyses with polyclonal anti-urease and anti-UreD antibodies indicated that UreD is masked in the UreD-UreF-urease complexes, presumably by UreF. We propose that the binding of UreF modulates the UreD-urease apoprotein activation properties by excluding nickel ions from binding to the active site until after formation of the carbamylated lysine metallocenter ligand.
Moncrief
M B
MB
Department of Biochemistry, Michigan State University, East Lansing 48824-1101, USA.
Hausinger
R P
RP
D001059:Apoproteins / Q000378:metabolism*; D001426:Bacterial Proteins / Q000302:isolation & purification / Q000378:metabolism*; D001639:Bicarbonates / Q000494:pharmacology; D004789:Enzyme Activation / Q000187:drug effects; D007711:Klebsiella pneumoniae / Q000201:enzymology*; D009532:Nickel / Q000494:pharmacology; D011485:Protein Binding; D014510:Urease / Q000302:isolation & purification / Q000378:metabolism*; D015032:Zinc / Q000494:pharmacology
D001059:Apoproteins; D001426:Bacterial Proteins; D001639:Bicarbonates; C086767:UreD protein, Bacteria; C101639:UreF protein, bacteria; D009532:Nickel; D014510:Urease; D015032:Zinc