Thiamine → L-Glutamic Acid

Participants

Reaction Steps

coli strains, VL334thrC+, VL334thrC+glpK*-231, VL334thrC+glpK*-234, VL334thrC+glpK*-random, and VL334thrC+glpK*-ΔglpR, can be grown for 18-24 hours at 37° C. on L-agar plates. Then, one loop of the cells can be transferred into test tubes containing 2 ml of fermentation medium. The fermentation medium contains glycerol (60 μl), ammonium sulfate (25 μl), KH2PO4 (2 g/l), MgSO4 (1 μl), thiamine (0.1 mg/ml), L-isoleucine (70 μg/ml), and CaCO3 (25 μl). The pH is adjusted to 7.2. Glyserol and CaCO3 are sterilized separately. Cultivation can be carried out at 30° C. for 3 days with shaking. After the cultivation, the amount of L-glutamic acid which is produced can be determined by paper chromatography (liquid phase composition of butanol-acetic acid-water=4:1:1) with subsequent staining by ninhydrin (1% solution in acetone) and further elution of the compounds in 50% ethanol with 0.5% CdCl2.

Provenance & Evidence