earticle

영어

Flavin-containing monooxygenases (FMOs) have been identified from a variety of organisms. Our previous studies found that recombinant E. coli cells harboring fmo gene from Methylophaga aminisulfidivorans MPT produced indigo (920 mg/L) and indirubin (≤5 mg/L) in a 5 L fermentor containing tryptophan medium (2g/L tryptophan, 10g/L NaCl and 5g/L yeast extract). Interestingly, indirubin production greatly increased when cysteine was added to the tryptophan medium. Although growth of the recombinant E. coli harboring fmo gene was inhibited by cysteine, protein expression level and activity of FMO were not influenced. The optimum culture conditions for indirubin production in tryptophan medium were as follows: 2 g/L tryptophan, 5 g/L yeast extract, 10 g/L NaCl, 3 mM cysteine, pH 8.0 and at 30℃. Under these conditions, recombinant E. coli cells produced 223.6 mg/L of indirubin. In order to investigate the proteins involved in indirubin production in E. coli, proteomic analysis was performed using two-dimensional gel electrophoresis in conjunction with MALDI-TOF mass spectrometry. The analysis showed that the expression levels of 66 proteins were varied quantitatively with statistical significance under the optimum indirubin production condition. Through classification of physiological functions from Matrix Science Database analysis, nine proteins, which maybe involved in tryptophan synthesis and biofilm formation, were identified.