Efficient in vivo synthesis and rapid purification of chorismic acid using an engineered Escherichia coli strain

Chorismate is a key intermediate in the biosynthesis of aromatic amino acids and other natural products in microorganisms and plants. Four chorismate mutase-deficient strains of Escherichia coli were evaluated for the large scale microbiological production of this metabolite which is needed for detailed biochemical and structural studies of chorismate-utilizing enzymes. Optimization of culturing and isolation conditions with one of these strains yielded an improved protocol for the production of chorismate in yields that compare favorably with those previously obtained with Klebsiella pneumoniae strain 62-1, a Class 2 pathogenic organism. A simple, single-step procedure for the purification of large quantities of chorismic acid by Cls reverse phase silica gel flash chromatography is also described. Chorismic acid obtained through this procedure is 90-98% pure and can be stored as the free acid in crystalline form over several months without decomposition. (C) 1997 Academic Press.