假单胞菌胞外酶降解黄曲霉毒素B1的酶学性质

Characterization of an extracellular aflatoxin B1 degrading enzyme from a Pseudomonas strain

摘要:

【背景】黄曲霉毒素B1 (Aflatoxin B1，AFB1)毒性强、污染普遍，目前尚无有效的防治办法。【目的】为了发掘高效的AFB1降解菌并探索其降解特性，对红树林污泥样品中一株AFB1降解菌株(HAI2)的酶学性质进行分析。【方法】以AFB1结构类似物为唯一碳源，筛选出一株高效的AFB1降解菌，利用16S rRNA基因测序结果鉴定菌株，并结合HPLC研究菌株对AFB1的降解特性。【结果】HAI2的16S rRNA基因序列与恶臭假单胞菌(Pseudomonas putida，NR 113651.1)的相似性高达99.85%，其主要降解AFB1的活性物质为胞外蛋白质，HAI2-AFB1降解酶的最适pH值为7.0，最适温度为37 °C，Fe2+、Ca2+和Zn2+会抑制AFB1的降解率，Cu2+和Mn2+可以提高AFB1的降解率。HAI2上清液对100 ng/mL的AFB1降解24 h的降解率为71.52%。此外，P. putida HAI2还可抑制被黄曲霉菌感染的玉米中AFB1的合成，AFB1含量可减少63.46%。【结论】假单胞菌P. putida HAI2具有较高的降解AFB1能力，其主要降解活性物质为胞外蛋白，在食品和饲料行业具有较大的应用前景。

关键词:黄曲霉毒素B1，筛选，假单胞菌，酶学性质，胞外蛋白

Abstract:

[Background] Aflatoxin B1 (aflatoxin B1, AFB1) is a highly toxic and widespread mycotoxin. At present, there is no effective method to control its pollution. [Objective] To discover efficient AFB1 degrading bacteria and explore its degradation characteristics, enzymatic properties of an AFB1 degrading strain (HAI2) from mangrove sludge samples were analyzed. [Methods] Using the structural analogue of AFB1 as the sole carbon source, a highly efficient AFB1 degrading strain was screened out, then 16S rRNA gene sequencing technology was used to identify the strain species, and HPLC was used to analyze the strain’s degradation characteristics to AFB1. [Results] 16S rRNA gene sequence of HAI2 shares 99.85% homology with Pseudomonas putida (NR 113651.1). The main component of degradation AFB1 was extracellular protein. According to the analysis of enzymatic properties, the optimum pH value of HAI2-AFB1 degrading enzyme was 7.0 and the optimum temperature was 37 °C. Fe2+, Ca2+, and Zn2+ could inhibit the degradation rate of AFB1, and Cu2+ and Mn2+ could increase the degradation rate of AFB1. The degradation rate of 100 ng/mL AFB1 by HAI2 supernatant was 71.52% for 24 h. Besides, P. putida HAI2 could inhibit AFB1 synthesis in corn infected by Aspergillus flavus, and the content could be reduced by 63.46%. [Conclusion] P. putida HAI2 had a high ability to degrade AFB1, and its main degradation active substance was extracellular protein, with a large application prospect in the food and feed industry.

Key words:Aflatoxin B1, screening, Pseudomonas, enzymatic properties, extracellular protein