常压室温等离子诱变选育L-天冬酰胺酶高产重组菌

Screening of High L-Asparaginase Activity Mutants of Recombinant Bacteria WB600 by Atmospheric and Room Temperature Plasmas Mutation System

以Bacillus subtilis WB600为宿主,构建了能分泌表达L-ASNase的重组菌,并通过常压室温等离子体诱变进一步提高了重组菌的产酶量。重组Bacillus subtilis WB600(p MA5-wap Aans Z)发酵30 h,胞外酶活达到37.2 U/m L,表明L-ASNase在信号肽wap A介导下能分泌至胞外。在功率120 W、气流量10 L/min、诱变时间40 s的诱变操作条件下,对重组菌进行了等离子体诱变。突变株的酶活最高达48.4 U/m L,较诱变前提高30%。上述结果表明,常压室温等离子体诱变能有效提高重组菌产L-ASNase的酶活.研究结果为L-ASNase的工业化生产提供了高效的生产菌株。

L-asparaginase（L-ASNase） can catalyze the deamidation of L-asparagine（a precursor of toxic acrylamide） to L-aspartic acid and ammonia,and it is thus an important food enzyme. This study used Bacillus subtilis WB600 as the expression host,successfully realizing the secretory expression of L-ASNase. By using the atmospheric and room temperature plasmas（ARTP） mutation system,the titer of the recombinant L-ASNase was improved further. After 30 h fermentation,extracellular enzyme activity of recombinant B. subtilis WB600（p MA5- wap A-ans Z）reached 37.2U/m L. This result suggested that L-ASNase can secrete to the medium mediated by a signal peptide of wap A. When treated with the ARTP at power input 120 W and gas flow rate 10 L/min for 40 s,a mutant of B. subtilis WB600（p MA5- wap A-ans Z）with a activity of 48.4 U/m L was obtained,the activity value was increased by 30%. These results suggested that ARTP mutation system can be used to improve the production of L-ASNase of recombinant strains. This study provided a good L-ASNase producing strain to fermentation production at an industrial scale.