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Cloning and Expression of β-mannanase Gene in Bacillus subtilis
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- Author:
- No author available
- Journal Title:
- Chinese Journal of Applied & Environmental Biology
- Issue:
- 4
- DOI:
- 10.3724/SP.J.1145.2012.00672
- Key Word:
- β-甘露聚糖酶;枯草芽孢杆菌;信号肽;克隆;表达;重组菌株;酶学性质
Abstract： 从Bacillus subtilis JNA 3-10中克隆出β-甘露聚糖酶基因成熟肽链编码序列manA1和含信号肽的β-甘露聚糖酶基因manA2,在B.subtilis 168中克隆表达,分别筛选获得高效分泌表达β-甘露聚糖酶的重组菌株BPM1001(pMA5-manAl/B.subtilis 168)和BPM1002(pMA5-manA2/B.subtilis 168),结果表明菌株BPM1002总酶活力是菌株BPM1001的9.65倍,是原始菌株的13.1倍.在基因manA2下游引入His序列克隆出β-甘露聚糖酶基因manA3,获得枯草芽孢杆菌168重组菌株BPM1003.采用Ni-NTA柱纯化重组菌株BPM1003分泌表达的β-甘露聚糖酶,并研究其酶学性质,该酶促反应的最适pH为6.5,最适温度为65℃,在37 ℃条件下保存一个月酶活力依然保留有77.8％.5 L发酵罐放大实验结果表明魔芋粉对于产β-甘露聚糖酶具有明显的诱导作用,酶活力最高可达2748.82 U/mL.
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