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.