Abstract

TdiA, a nonribosomal peptide synthetase, catalyzes the carbon-carbon bond formation in biosynthesis of bis-indolylquinone natural products. The TdiA gene (tdiA) containing several codons rarely used by Escherichia coli, was cloned from the genome ofAspergillus nidulans and optimized in two strains of E. coli: BL21 (DE3) and Rosetta (DE3), which is a rare codon optimizer strain. The effects of initial isopropyl-β-D-thiogalactopyranoside (IPTG) concentration and induction time on the enzyme expression level were investigated in two strains. The results indicated that the amount of TdiA expressed in Rosetta (DE3) was about 6-fold higher than that in BL21 (DE3). The activity of TdiA expressed by Rosetta (DE3) in enzymatic synthesis of bis-indolylquinone using indole-3-pyruvic acid (IPA) as substrate was generally the same as that expressed in BL21 (DE3). Based on the optimal culture system using Rosetta (DE3), the yield of TdiA achieved 10.32 mg/L under the appropriate conditions. This efficient expression of TdiA would be in favour of advancing the totally enzymatic preparation of bis-indolylquinone natural products.

Key words: Nonribosomal peptide synthetase, Aspergillus nidulans, Escherichia coliRosetta (DE3), rare codons, bis-indolylquinone.