节杆菌HW08降解苦马豆素关键基因在乳酸乳球菌中的表达及降解功能分析

Expression and degradation function analysis of the key genes from Arthrobacter nitroquajacolicus HW08 in Lactococcus lactis for the degradation of swainsonine

摘要:

为缓解疯草中毒病对我国畜牧业的不良影响并充分利用潜在牧草资源，利用生物降解技术对疯草进行处理，可以去除疯草中的主要毒性成分苦马豆素，使之作为优质牧草被利用。节杆菌HW08被证实对苦马豆素具有稳定高效的降解能力。本研究以被公认为安全的食品级微生物的乳酸乳球菌作为表达载体，选取节杆菌HW08中的4个关键降解基因进行表达，并使用液相色谱法对其降解苦马豆素的能力进行了分析。结果显示，转化了乙醇脱氢酶A1R6C3基因的乳酸乳球菌经诱导后，提取的粗酶液在30℃条件下，24 h内能够降解约323.4 μg苦马豆素；转化了谷胱甘肽合酶、酯酶/酰基水解酶、糖基转移酶基因的乳酸乳球菌中提取的粗酶液单独作用时未表现出对于苦马豆素的降解能力，但混合后在同样条件下能够降解约140.5 μg苦马豆素。本研究为苦马豆素降解工程菌的临床推广提供了帮助，也为动物苦马豆素中毒的防治及疯草的脱毒利用提供了新的研究思路。

Abstract:

In order to mitigate the adverse effects of madrassa poisoning disease on our livestock industry and to fully utilize the potential pasture resources, biodegradation of locoweed can remove swainsonine, the major toxic component of locoweed, so that the locoweed can be used as high-quality forage. Arthrobacter nitroquajacolicus HW08 can stably and efficiently degrade swainsonine. In this study, Lactococcus lactis, as a food-grade microorganism, was used as a vector to express four key degradation genes from A. nitroquajacolicus HW08. Subsequently, liquid chromatography was employed to evaluate the swainsonine-degrading performance. The crude enzyme solution extracted from the L. lactis strain transformed with the ethanol dehydrogenase gene A1R6C3 degraded 323.4 μg of swainsonine in 24 h at 30 ℃. The crude enzyme solutions from the L. lactis strains transformed with the genes encoding glutathione synthase, esterase/acyl hydrolase, and glycosyltransferase did not show any degradation ability for swainsonine when being used alone but degraded about 140.5 μg of swainsonine when being used in mixture. The findings will help the clinical promotion of swainsonine-degrading engineering strains and provide new research ideas for the prevention and treatment of swainsonine poisoning in animals and the detoxification and utilization of locoweed.