THERMOSTABLE D-HYDANTOINASE FROM THERMOPHILIC BACILLUS-STEAROTHERMOPHILUS SD-1 - CHARACTERISTICS OF PURIFIED ENZYME

One thousand thermophiles isolated from soils were screened for hydant oinase and its thermostability. One thermophilic bacterium that showed the highest thermostability and activity of hydantoinase was identifi ed to be Bacillus stearothermophilus SD-1 according to morphological a nd physiological characteristics, The hydantoinase of B. stearothermop hilus SD-1 was purified to homogeneity via ammonium sulfate fractionat ion, anion-exchange chromatography, heat treatment: hydrophobic-intera ction chromatography and preparative gel electrophoresis. The relative molecular mass of the hydantoinase was determined to be 126 kDa by ge l-filtration chromatography, and a value of 54 kDa was obtained as a m olecular mass of the subunit on analytical sodiumdodecylsulfate/polyac rylamide gel electrophoresis. The hydantoinase was strictly D-specific and metal-dependent. The optimal pH and temperature were about 8.0 an d 65 degrees C respectively, and the half-life of the D-hydantoinase w as estimated to be 30 min at 80 degrees C, indicating the most thermos table enzyme so far.