Purification and characterization of urease from Ureaplasma urealyticum.
The urease from Ureaplasma urealyticum strain T 960 was isolated by the use of affinity chromatography, hydrophobic chromatography and gel filtration. The enzyme was purified by a factor of 155. The urease appeared as a single band of molecular weight (MW) 75,000 using reducing conditions in SDS-polyacrylamide gel electrophoresis. By gel filtration the native MW was determined to be 150,000. Isoelectric focusing showed the presence of two closely migrating enzyme species with a pI of pH 5.1-5.2. These findings show multiple forms of the urease and that these forms are composed of subunits. The electrophoresis experiments also indicate that this enzyme is a major component of the cytoplasm of U. urealyticum. The Km of the purified enzyme was 4.5 mM urea and the specific activity was 33530 mumoles NH3 x min-1 x mg-1. The optimum pH was pH 7-7.5. The urease activity was inhibited by flurofamide, acetohydroxamic acid, N-ethylmaleimide and p-chloromercuribenzoate but not by iodoacetate.
Saada
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Department of Membrane and Ultrastructure Research, Hebrew University-Hadassah Medical School, Jerusalem, Israel.
Kahane
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D001549:Benzamides / Q000494:pharmacology; D002729:Chloromercuribenzoates / Q000494:pharmacology; D002846:Chromatography, Affinity; D002847:Chromatography, Agarose; D002850:Chromatography, Gel; D004591:Electrophoresis, Polyacrylamide Gel; D005033:Ethylmaleimide / Q000494:pharmacology; D006868:Hydrolysis; D006877:Hydroxamic Acids / Q000494:pharmacology; D007461:Iodoacetates / Q000494:pharmacology; D007525:Isoelectric Focusing; D008970:Molecular Weight; D014508:Urea / Q000378:metabolism; D014509:Ureaplasma / Q000201:enzymology*; D014510:Urease / Q000032:analysis / Q000037:antagonists & inhibitors / Q000302:isolation & purification* / Q000378:metabolism
D001549:Benzamides; D002729:Chloromercuribenzoates; D006877:Hydroxamic Acids; D007461:Iodoacetates; C006358:acetohydroxamic acid; C032764:flurofamide; D014508:Urea; D014510:Urease; D005033:Ethylmaleimide