AMPLIFICATION OF THE TRYPTOPHAN OPERON GENE IN ESCHERICHIA-COLI CHROMOSOME TO INCREASE L-TRYPTOPHAN BIOSYNTHESIS

Classical mutagenesis could desensitize the feedback inhibition of L-t ryptophan (L-Trp) biosynthesis. Among the mutants, a 5-fluorotryptopha n-resistant strain, Escherichia coli EMS4-C25 produced 3 g/l of L-Trp within 18 h. The feedback-resistant L-Trp operon gene (trp) prepared f rom E. coli EMS4-C25 was inserted into pUC19 and pHSG576 to generate p TC701 and pTC576, respectively. When pHSG576 and pTC701 were introduce d into E. coli EMS4-C25, chromosomal integration occurred through homo logous recombination. By using Southern hybridization, we demonstrated that the integrated plasmids existed as multicopies. The strains with integrated foreign trp operon gene had higher activities of anthranil ate synthase and Trp synthase than those found for the host strain and produced 9.2 g/l of L-Trp with 13% converison yield from D-glucose. T he integration and amplification of the trp-operon-bearing plasmid avo ided the plasmid instability and increased L-Trp production.