초록
<P><B>Background</B></P><P>L-tryptophan is an aromatic amino acid widely used in the food, chemical and pharmaceutical industries. In <I>Escherichia coli</I>, L-tryptophan is synthesized from phosphoenolpyruvate and erythrose 4-phosphate by enzymes in the shikimate pathway and L-tryptophan branch pathway, while L-serine and phosphoribosylpyrophosphate are also involved in L-tryptophan synthesis. In order to construct a microbial strain for efficient L-tryptophan production from glucose, we developed a one step tryptophan attenuator inactivation and promoter swapping strategy for metabolic flux optimization after a base strain was obtained by overexpressing the <I>tktA</I>, mutated <I>trpE </I>and <I>aroG </I>genes and inactivating a series of competitive steps.</P><P><B>Results</B></P><P>The engineered <I>E. coli </I>GPT1002 with tryptophan attenuator inactivation and tryptophan operon promoter substitution exhibited 1.67 ~ 9.29 times higher transcription of tryptophan operon genes than the control GPT1001. In addition, this strain accumulated 1.70 g l<SUP>-1 </SUP>L-tryptophan after 36 h batch cultivation in 300-mL shake flask. Bioreactor fermentation experiments showed that GPT1002 could produce 10.15 g l<SUP>-1 </SUP>L-tryptophan in 48 h.</P><P><B>Conclusions</B></P><P>The one step inactivating and promoter swapping is an efficient method for metabolic engineering. This method can also be applied in other bacteria.</P>