Arcand N, Kluepfel D, Paradis F W, Morosoli R, Shareck F
Centre de Recherche en Microbiologie Appliquée, Institut Armand-Frappier, Université du Québec, Laval-des-Rapides, Canada.
Biochem J. 1993 Mar 15;290 ( Pt 3)(Pt 3):857-63. doi: 10.1042/bj2900857.
The gene coding for a beta-mannanase was cloned homologously from Streptomyces lividans and its DNA sequence was determined. The fully secreted enzyme was isolated and purified from culture filtrates of the hyperproducing clone S. lividans IAF36 grown in mineral salt media containing galactomannan as the main carbon source. It had a molecular mass of 36 kDa and a specific activity of 876 i.u./mg of protein. Under the assay conditions used, the optimal enzyme activity was obtained at 58 degrees C and a pH of 6.8. The pI was 3.5. The kinetic constants of this mannanase determined with galactomannan as substrate were a Vmax. of 205 i.u./mg of enzyme and a Km of 0.77 mg/ml. Data from SDS/PAGE and Western blotting show that the cloned enzyme was identical to that of the wild-type strain.
从淡紫链霉菌中同源克隆了编码β-甘露聚糖酶的基因，并测定了其DNA序列。从以半乳甘露聚糖作为主要碳源的无机盐培养基中生长的高产克隆淡紫链霉菌IAF36的培养滤液中分离并纯化了完全分泌的酶。它的分子量为36 kDa，比活性为876 i.u./mg蛋白质。在所使用的测定条件下，最佳酶活性在58℃和pH 6.8时获得。其等电点为3.5。以半乳甘露聚糖为底物测定的该甘露聚糖酶的动力学常数为：Vmax为205 i.u./mg酶，Km为0.77 mg/ml。SDS/PAGE和蛋白质印迹数据表明，克隆的酶与野生型菌株的酶相同。
