Phillips A, Pretorius G H, Du Toit P J
Department of Microbiology and Biochemistry, University of the Orange Free State, Bloemfontein, Republic of South Africa.
FEMS Microbiol Lett. 1991 Mar 15;63(1):21-5. doi: 10.1016/0378-1097(91)90520-k.
Cell-free extracts of a selection of yeasts were analysed for urease activity. Species in the genera Filobasidiella, Rhodotorula and Rhodosporidium had the highest specific activities. Immune inactivation experiments showed widely different degrees of cross-reactivity between antiserum to jack bean urease and yeast ureases, with Rhodosporidium paludigenum (71%) the most and Schizosaccharomyces pombe (3%) the least affected. Only R. paludigenum urease was detected with anti-jack bean urease antiserum on Western blots. The urease of Rhodosporidium paludigenum was partially purified by column chromatography. The native enzyme was found to have a subunit size of 72 +/- 7 kDa probably in an octamer arrangement of 560 +/- 8 kDa, having a specific activity of 62.5 mumol urea hydrolysed min-1 (mg protein)-1. The enzyme was stable in the pH range 5-11 with optimum activity at pH 7.8. Vmax and Km values were determined as 65.2 +/- 3.8 mumol min-1 (mg protein)-1 and 3.81 +/- 0.47 mM, respectively.
对一系列酵母的无细胞提取物进行了脲酶活性分析。丝孢酵母属、红酵母属和红冬孢酵母属中的物种具有最高的比活性。免疫失活实验表明，刀豆脲酶抗血清与酵母脲酶之间的交叉反应程度差异很大，其中沼泽红冬孢酵母（71%）受影响最大，粟酒裂殖酵母（3%）受影响最小。在蛋白质印迹法中，只有沼泽红冬孢酵母脲酶能被刀豆脲酶抗血清检测到。通过柱色谱法对沼泽红冬孢酵母脲酶进行了部分纯化。发现天然酶的亚基大小为72±7 kDa，可能以560±8 kDa的八聚体形式存在，比活性为62.5 μmol尿素每分钟水解量每毫克蛋白质。该酶在pH 5 - 11范围内稳定，在pH 7.8时活性最佳。Vmax和Km值分别测定为65.2±3.8 μmol每分钟每毫克蛋白质和3.81±0.47 mM。
