title: Development of indole-3-acetic acid-producing Escherichia coli by functional expression of IpdC, AspC, and Iad1.
abstract: Biosynthesis of indole-3-acetic acid (IAA) via the indole-3-pyruvic acid pathway involves three kinds of enzymes; aminotransferase encoded by aspC, indole-3-pyruvic acid decarboxylase encoded by ipdC, and indole-3-acetic acid dehydrogenase encoded by iad1. The ipdC from Enterobacter cloacae ATCC 13047, aspC from Escherichia coli, and iad1 from Ustilago maydis were cloned and expressed under the control of the tac and sod promoters in E. coli. According to SDS-PAGE and enzyme activity, IpdC and Iad1 showed good expression under the control of P(tac), whereas AspC was efficiently expressed by P(sod) originating from Corynebacterium glutamicum. The activities of IpdC, AspC, and Iad1 from the crude extracts of recombinant E. coli Top 10 were 215.6, 5.7, and 272.1 nmol/min/mg-protein, respectively. The recombinant E. coli DH5α expressing IpdC, AspC, and Iad1 produced about 1.1 g/l of IAA and 0.13 g/l of tryptophol (TOL) after 48 h of cultivation in LB medium with 2 g/l tryptophan. To improve IAA production, a tnaA gene mediating indole formation from tryptophan was deleted. As a result, E. coli IAA68 with expression of the three genes produced 1.8 g/l of IAA, which is a 1.6- fold increase compared with wild-type DH5α harboring the same plasmids. Moreover, the complete conversion of tryptophan to IAA was achieved by E. coli IAA68. Finally, E. coli IAA68 produced 3.0 g/l of IAA after 24 h cultivation in LB medium supplemented with 4 g/l of tryptophan.
authors.affiliation: Department of Food Science and Biotechnology, Kyungsung University, Busan 608-736, Republic of Korea.
authors.forename: Elisa Friska
authors.initials: EF
authors.lastname: Romasi
authors.forename: Jinho
authors.initials: J
authors.lastname: Lee
mesh_terms: D053898:Biosynthetic Pathways / Q000235:genetics*; D003001:Cloning, Molecular; D048230:Corynebacterium glutamicum / Q000235:genetics; D016972:Enterobacter cloacae / Q000201:enzymology / Q000235:genetics; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics* / Q000378:metabolism*; D015870:Gene Expression; D007210:Indoleacetic Acids / Q000378:metabolism*; D060847:Metabolic Engineering / Q000379:methods*; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D014588:Ustilago / Q000201:enzymology / Q000235:genetics
chemical_list: D007210:Indoleacetic Acids; D011994:Recombinant Proteins; C030737:indoleacetic acid