摘要
为提高阿魏酸酯酶(ferulic esterase,FAE)的发酵单位活力,将土曲霉FAE编码基因在毕赤酵母实现异源高效表达。重组毕赤酵母发酵活力为(17.38±0.34)U/mL,是原始菌株的35倍左右。经十二烷基硫酸钠-聚丙烯酰胺凝胶电泳检验得出,重组土曲霉阿魏酸酯酶(recombinant Aspergillus terreus ferulic esterase,rAtFAE)分子质量约为39 kDa。酶学性质研究表明rAtFAE最适pH值为6.0、最适反应温度为50℃,以咖啡酸甲酯、香豆酸甲酯、阿魏酸甲酯、芥子酸甲酯为底物时测得的kcat/Km值分别为255、237、112.64、96.85 L/(mol·min)。在以麸皮为原料制备阿魏酸的应用实验中,rAtFAE可以与木聚糖酶产生良好的协同作用并有效提升阿魏酸的释放量,最大释放量为(1 876.45±30.05)μg/g(以麸皮质量计)。
To improve the expression level of ferulic esterase(FAE),the FAE encoding gene from Aspergillus terreus was heterologously expressed in Pichia pastoris.The expression level of FAE in the recombinant strain was(17.38±0.34)U/mL,which was approximately 35 times as high as that of the original strain.The molecular mass of the recombinant A.terreus FAE(rAtFAE)was approximately 39 kDa as estimated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis(SDS-PAGE).Enzymatic characterization showed that the optimal pH of rAtFAE was 6.0,the optimal reaction temperature was 50℃,and the kcat/Km values measured with methyl caffeic acid,methyl coumarinate,methyl ferulic acid and methyl sinakoate as substrates were 255,237,112.64 and 96.85 L/(mol·min),respectively.The combination of rAtFAE and xylanase showed a good synergistic effect in enhancing the release of ferulic acid from wheat bran up to(1876.45±30.05)μg/g.
作者
韩红祥
郭成城
魏胜华
陈阿娜
李松
HAN Hongxiang;GUO Chengcheng;WEI Shenghua;CHEN Ana;LI Song(School of Biological and Food Engineering,Anhui Polytechnic University,Wuhu 241000,China)
出处
《食品科学》
北大核心
2025年第15期155-163,共9页
Food Science
基金
安徽高校自然科学研究项目(KJ2021A0509)
安徽省自然科学基金项目(2208085MC83)。
关键词
土曲霉
阿魏酸酯酶
异源表达
毕赤酵母
阿魏酸
Aspergillus terreus
ferulic esterase
heterologous expression
Pichia pastoris
ferulic acid
