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

摘要:

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（pMA5- wapA -ansZ）发酵30 h，胞外酶活达到37.2 U/mL，表明L-ASNase在信号肽wapA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/mL，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。

Abstract:

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（pMA5- wapA -ansZ）reached 37.2 U/mL. This result suggested that L-ASNase can secrete to the medium mediated by a signal peptide of wapA. 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（pMA5- wapA -ansZ）with a activity of 48.4 U/mL 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.