基于重组大肠杆菌无细胞体系生产吡咯喹啉醌

摘要

采用无细胞体系生产吡咯喹啉醌（pyrroloquinoline quinone，PQQ）。首先将肺炎克雷伯氏菌PQQ基因簇pqqABC-DEF置于半乳糖苷酶启动子之下，构建表达载体，经转化筛选获得重组大肠杆菌。制备重组菌的细胞匀浆，体外反应后测定PQQ产量。结果显示，与活体重组菌相比，无细胞体系的PQQ产量提高约30%，表明胞内存在PQQ合成的限速反应，而无细胞体系可解除此限速反应。%In this study, cell-free system was developed to overproduce pyrroloquinoline quinone(PQQ). The PQQ gene cluster from Klebsiella pneumoniae was subcloned into vector harboring lac promoter and a recombinant Escherichia coli was acquired. Subsequently cell homogenate was prepared which showed~30%increase of PQQ yield compared with in vivo biosynthesis. Not only revealing the presence of intracellular velocity-limiting reaction(s)that retards PQQ biosynthesis, this study also suggests that cell-free system can address this issue. Hence, this study provides basis for further enhancement of PQQ production.