PQQ依赖性呕吐毒素脱氢酶的鉴定与性能表征  

Identification and Biochemical Characterization of a PQQ-dependent Deoxynivalenol(DON)Dehydrogenase

作　　者：汪洋 赵冬蕾 王楠希 杨永坛 郭宝元 WANG Yang;ZHAO Dong-lei;WANG Nan-xi;YANG Yong-tan;GUO Bao-yuan(Institute of Grain and Oil Quality and Safety,Academy of National Food and Strategic Reserves Administration,Beijing 100037,China;School of Health Science and Engineering,University of Shanghai for Science and Technology,Shanghai 200093,China)

机构地区：[1]国家粮食和物资储备局科学研究院粮油质量安全研究所,北京100037 [2]上海理工大学健康科学与工程学院,上海200093

出　　处：《粮油食品科技》2023年第5期179-187,共9页Science and Technology of Cereals,Oils and Foods

基　　金：国家自然科学基金(31972605);宁夏回族自治区重点研发计划(2022BBF03031)。

摘　　要：谷物及其制品中呕吐毒素(DON)的污染给人类健康造成了潜在威胁。利用生物酶将真菌毒素转化为低毒产物的生物脱毒技术是一种环保且高效的脱毒方法。本研究基于同源比对方法,从呕吐毒素脱毒菌株德沃斯氏菌Devosia sp.FJ2-5-3的基因组中挖掘鉴定了1个呕吐毒素(DON)脱毒酶编码基因adh2,对该基因进行克隆、表达并进一步对其编码酶的酶学性质进行了表征。结果表明:adh2基因长度1770 bp,其编码的ADH2酶原由589个氨基酸组成,N端前1~24个氨基酸为信号肽,属于Ⅰ型吡咯喹啉醌(Pyrroloquinoline quinone,PQQ)依赖性醇脱氢酶。该酶最适反应pH=9,最适反应温度为35℃,在65℃孵育4 h后仍保留50%以上的酶活,显示了较好的热稳定性;另外,Ca^(2+)是其必须辅因子,而1%的乙醇和异丙醇均对其活性有明显抑制作用。ADH2对DON的Km和Kcat值分别为(708.00±47.01)μg/mL和(3.37±0.11)S^(-1),该研究结果可为PQQ依赖性呕吐毒素脱氢酶后续工业化应用提供参考。Deoxynivalenol(DON)contamination in cereals and cereal products has the potential to pose a threat to human health.The biodetoxification technique,which uses biological enzymes to transform mycotoxins into less toxic product,is an environmentally friendly and highly effective approach to detoxifying mycotoxins.In this study,a gene designated adh2,encoding DON detoxifying enzyme,was identified by homology search from the genome sequence of DON-detoxifying strain Devosia sp.FJ2-5-3.Subsequently.The adh2 gene was cloned and expressed.Then,the enzymatic properties of its encoding enzyme was further characterized.The results showed that the length of the adh2 gene in Devosia sp.FJ2-5-3 was 1770 bp and the encoded ADH2 enzyme consisted of 589 amino acids,with a signal peptide of 24 amino acids length present at the N-terminus and belonging to type I of the PQQ-dependent alcohol dehydrogenase family(Type I PQQ-ADH).The enzyme exhibited the optimal activity at pH 9 and a temperature of 35°C,and it retained over 50%of enzyme activity after incubation at 65℃for 4 hours,indicating its good thermal stability.In addition,Ca^(2+)was found to be an essential cofactor for ADH2,while both 1%of ethanol and isopropanol showed significant inhibitory effects on its activity.The K_(m)and K_(cat)values of ADH2 for DON were 708.00±47.01μg/mL and 3.37±0.11 S^(-1),respectively.These results could provide reference information for the subsequent industrial application of DON detoxification enzymes.

关 键 词：呕吐毒素 生物脱毒 PQQ依赖性醇脱氢酶 酶学性质表征 

分 类 号：TS207[轻工技术与工程—食品科学] Q93[轻工技术与工程—食品科学与工程]