Microbial synthesis of p-hydroxybenzoic acid from glucose

A series of recombinant Escherichia coli strains have been constructed and evaluated for their ability to synthesize p-hydroxybenzoic acid from glucos e under fed-batch fermentor conditions. The maximum concentration of p-hydr oxybenzoic acid synthesized was 12 g/L and corresponded to a yield of 13% ( mol/mol). Synthesis of p-hydroxybenzoic acid began with direction of increa sed carbon flow into the common pathway of aromatic amino acid biosynthesis . This was accomplished in all constructs with overexpression of a feedback -insensitive isozyme of 3-deoxy-D-arabino-heptulosonic acid 7-phosphate syn thase. Expression levels of enzymes fn the common pathway of aromatic amino acid biosynthesis were also increased in all constructs to deliver increas ed carbon flow from the beginning to the end of the common pathway. A previ ously unreported inhibition of 3-dehydroquinate synthase by L-tyrosine was discovered to be a significant impediment to the flow of carbon through the common pathway. Chorismic acid, the last metabolite of the common pathway, was converted into p-hydroxybenzoic acid by ubiC-encoded chorismate lyase. Constructs differed in the strategy used for overexpression of chorismate lyase and also differed as: to whether mutations were present in the host E colt to inactivate other chorismate-utilizing enzymes. Use of overexpresse d chorismate lyase to increase the rate of chorismic acid aromatization was mitigated by attendant decreases in the specific activity of DAHP synthase and feedback inhibition caused by p-hydroxybenzoic acid. The toxicity of p -hydroxybenzoic acid towards E. coli metabolism and growth was also evaluat ed. (C) 2001 John Wiley & Sons, Inc.