Abstract:

The gene manS221 encoding a β-l, 4-mananase was cloned from Streptomyces fradiae var. k11 by degenerate primers and TAIL-PCR technology. manS221 gene contains 1 359 bp, encoding a 452 amino acid peptide consisting of a putative N-terminal signal peptide of 28 residues. The expression vector pET-28a-manS221 was constructed and transformed into Eschetichia coli BL21. The expressed ManS221 was purified by Ni-NTA chromatography. The optimum temperature and pH of the recombinant mannose were 52℃ and 6.0, respectively. The enzyme showed stability over a pH range of 5.0~9.0, and resistance to neutral protease. These favorable properties would make ManS221 as a good candidate in various industrial applications.

Key words: Streptomyces fradiae var. k11, mannose, manS221 gene, protease resistance

摘要：

通过设计简并引物和TAIL-PCR的方法从Streptomyces fradiae var. k11中克隆得到一个β-l,4-甘露聚糖酶编码基因manS221,其全长1 359 bp,编码452个氨基酸,前端28个氨基酸为预测信号肽。构建表达载体pET-28a-manS221并转化大肠杆菌BL2l（DE3）,特异性表达manS221基因, 重组的甘露聚糖酶通过Ni-NTA亲和层析纯化。酶学性质分析表明,重组甘露聚糖酶最适温度为52℃,最适pH值为6.0,在pH 5.0~9.0的范围内有良好的稳定性,并且对中性蛋白酶具有高抗性。这些优良的性质使得ManS221在食品、饲料和纺织工业生产中具有良好的应用前景。

关键词: 链霉菌Streptomyces fradiae var. k11;甘露聚糖酶;manS221基因；蛋白酶抗性