5-Hydroxytryptophan (5-HTP) is the precursor of the neurotransmitter serotonin and has been used for the treatment of various diseases such as depression, insomnia, chronic headaches, and binge eating associated obesity. The production of 5-HTP had been achieved in our previous report, by the development of a recombinant strain containing two plasmids for biosynthesis of L-tryptophan (L-trp) and subsequent hydroxylation. In this study, the L-trp biosynthetic pathway was further integrated into the E. coli genome, and the promoter strength of 3-deoxy-7-phosphoheptulonate synthase, which catalyzes the first step of L-trp biosynthesis, was engineered to increase the production of L-trp. Hence, the 5-HTP production could be manipulated by the regulation of copy number of L-trp hydroxylation plasmid. Finally, the 5-HTP production was increased to 1.61 g/L in the shaking flasks, which was 24% improvement comparing to the original producing strain, while the content of residual L-trp was successfully reduced from 1.66 to 0.2 g/L, which is beneficial for the downstream separation and purification. Our work shall promote feasible progresses for the industrial production of 5-HTP.