The urease of Ureaplasma urealyticum.
The urease from Ureaplasma urealyticum (serotype 8) has been purified by immuno-affinity column chromatography. Two active nickel-containing forms of the enzyme were demonstrated by non-denaturing electrophoretic analysis and a single active peak of apparent molecular mass 190 kDa was shown by FPLC. Total inactivation and denaturation of the enzyme to give three subunit polypeptides (one of 72 kDa containing nickel, one of 14 kDa and one of 11 kDa) was achieved by treatment with SDS and boiling. Densitometry suggested that the active enzyme contains equimolar ratios of the three subunits and hence is a hexamer. The enzyme displayed a pH optimum of 6.9 and pI values were determined. Storage of the purified enzyme at -70 degrees C followed by thawing to 20 degrees C caused a partial breakdown to inactive subunits. Anti-urease monoclonal antibodies bound both to the active enzyme and to the inactive 72 kDa subunit, and the antibodies cross-reacted with ureases from all of the other human serotypes. Competition assays with the antibodies revealed four distinct epitopes of the enzyme, all distinct from its active site.
Thirkell
D
D
Department of Biochemistry and Microbiology, University of St Andrews, Fife, UK.
Myles
A D
AD
Precious
B L
BL
Frost
J S
JS
Woodall
J C
JC
Burdon
M G
MG
Russell
W C
WC
D000911:Antibodies, Monoclonal / Q000276:immunology; D002846:Chromatography, Affinity; D000939:Epitopes / Q000032:analysis; D009532:Nickel / Q000032:analysis; D010455:Peptides / Q000032:analysis; D014509:Ureaplasma / Q000201:enzymology*; D014510:Urease / Q000032:analysis / Q000276:immunology / Q000302:isolation & purification*
D000911:Antibodies, Monoclonal; D000939:Epitopes; D010455:Peptides; D009532:Nickel; D014510:Urease