Pisani F M, Rella R, Raia C A, Rozzo C, Nucci R, Gambacorta A, De Rosa M, Rossi M
Istituto di Biochimica delle Proteine ed Enzimologia, Consiglio Nazionale delle Ricerche, Napoli, Italia.
Eur J Biochem. 1990 Jan 26;187(2):321-8. doi: 10.1111/j.1432-1033.1990.tb15308.x.
A thermophilic and thermostable beta-galactosidase activity was purified to homogeneity from crude extracts of the archaebacterium Sulfolobus solfataricus, by a procedure including ion-exchange and affinity chromatography. The homogeneous enzyme had a specific activity of 116.4 units/mg at 75 degrees C with o-nitrophenyl beta-galactopyranoside as substrate. Molecular mass studies demonstrated that the S. solfataricus beta-galactosidase was a tetramer of 240 +/- 8 kDa composed of similar or identical subunits. Comparison of the amino acid composition of beta-galactosidase from S. solfataricus with that from Escherichia coli revealed a lower cysteine content and a lower Arg/Lys ratio in the thermophilic enzyme. A rabbit serum, raised against the homogeneous enzyme did not cross-react with beta-galactosidase from E. coli. The enzyme, characterized for its reaction requirements and kinetic properties, showed a thermostability and thermophilicity notably greater than those reported for beta-galactosidases from other mesophilic and thermophilic sources.
通过包括离子交换和亲和色谱的方法，从嗜热古细菌嗜热栖热菌的粗提物中纯化出一种嗜热且耐热的β-半乳糖苷酶活性至均一。该均一酶以邻硝基苯基β-吡喃半乳糖苷为底物，在75℃时比活性为116.4单位/毫克。分子量研究表明，嗜热栖热菌β-半乳糖苷酶是由相似或相同亚基组成的240±8 kDa的四聚体。嗜热栖热菌β-半乳糖苷酶与大肠杆菌β-半乳糖苷酶的氨基酸组成比较显示，嗜热酶中的半胱氨酸含量较低，精氨酸/赖氨酸比率也较低。用该均一酶免疫家兔制备的血清与大肠杆菌的β-半乳糖苷酶不发生交叉反应。该酶经反应条件和动力学性质表征，显示出的耐热性和嗜热性明显高于其他中温及嗜热来源的β-半乳糖苷酶。
