九州大学大学院農学研究科遺伝子資源工学専攻遺伝子制御学講座
Laboratory of Molecular Gene Technics, Graduate School of Genetic Resources Technology, Faculty of Agriculture, Kyushu University
九州大学大学院農学研究科遺伝子資源工学専攻遺伝子制御学講座
Laboratory of Molecular Gene Technics, Graduate School of Genetic Resources Technology, Faculty of Agriculture, Kyushu University
九州大学大学院農学研究科遺伝子資源工学専攻遺伝子制御学講座
Laboratory of Molecular Gene Technics, Graduate School of Genetic Resources Technology, Faculty of Agriculture, Kyushu University
九州大学大学院農学研究科遺伝子資源工学専攻遺伝子制御学講座
Laboratory of Molecular Gene Technics, Graduate School of Genetic Resources Technology, Faculty of Agriculture, Kyushu University
To elucidate the expression mechanism and to attempt application of Streptomyces ATP nucleotide 3'-pyrophosphokinase (PPKase) gene in heterologous hosts, we expressed PPKase gene in E. coli cells. This was carried out by inserting a DNA fragment including PPKase gene downstream of the lac promoter in a multi-copy plasmid pUC119. The resultant plasmid, pUP3 was introduced into E. coli JM109. When the cells were incubated in the presence of IPTG, PPKase protein was found to be expressed and secreted into the periplasmic space. PPKase promoter was not functional in E. coli like most Streptomyces promoters. The increased activity of PPKase in pUP317, a deletion plasmid from pUP3, indicated that 5'-flanking sequence was very important to express PPKase gene. Various 3'-pyrophosphoryl nucleotides were detected, and growth inhibition was observed in PPKase-producing E. coli cells. Further studies are necessary for practical application of PPKase and its gene.
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