College of Bioengineering, Sichuan University of Science and Engineering, Yibin, Sichuan, China.
Guangdong Vtr Bio-Tech Co., Ltd., Zhuhai, Guangdong, China.
Biotechnol J. 2024 Jan;19(1):e2300167. doi: 10.1002/biot.202300167. Epub 2023 Dec 13.
In this study, three aflatoxin degrading enzyme genes, tv-adtz, arm-adtz and cu-adtz, were heterologously expressed in Pichia pastoris. The protein expression of the enzyme solution was detected by sodium dodecyl sulfate polyacrylamide gel electrophoresis, and the results showed that specific protein bands were detected and the target genes were successfully integrated into Pichia pastoris. The enzyme activities and detoxification efficiency of TV-ADTZ, Arm-ADTZ and Cu-ADTZ crude enzyme solutions were detected, and the highest enzyme activities were up to 3.57, 4.30, and 2.41 U mL , and the highest degradation rates were up to 45.58%, 60.0% and 34.21%, respectively. Arm-ADTZ with the best degradation effect was selected and designed for detoxification application experiments to test its detoxification efficiency of AFB in aqueous phase and in the process of moldy ground corn and preparation of DDGS, respectively, and the degradation rates reached 78.94%, 56.48%, and 24.31% after 24 h of reaction, respectively. Thus, it can be seen that the aflatoxin-degrading enzyme gene was successfully integrated into Pichia pastoris and secreted for expression, and the expressed product could effectively degrade AFB .
在这项研究中，三种黄曲霉毒素降解酶基因 tv-adtz、arm-adtz 和 cu-adtz 在巴斯德毕赤酵母中进行了异源表达。通过十二烷基硫酸钠聚丙烯酰胺凝胶电泳检测酶溶液的蛋白表达，结果表明检测到了特异性蛋白条带，并且目的基因成功整合到了巴斯德毕赤酵母中。检测了 TV-ADTZ、Arm-ADTZ 和 Cu-ADTZ 粗酶溶液的酶活性和解毒效率，最高酶活分别达到 3.57、4.30 和 2.41 U/mL，最高降解率分别达到 45.58%、60.0%和 34.21%。选择具有最佳降解效果的 Arm-ADTZ 进行解毒应用实验，分别测试其在水相、霉变玉米和 DDGS 制备过程中对 AFB 的解毒效率，反应 24 小时后，降解率分别达到 78.94%、56.48%和 24.31%。因此，可以看出黄曲霉毒素降解酶基因已成功整合到巴斯德毕赤酵母中并进行了分泌表达，表达产物能有效降解 AFB。
