Excessive acetic acid is harmful to tryptophan fermentation, and thus the level of acetic acid is normally restricted through the regulation of glucose content in the broth. In this study, the tdcD gene in Escherichia coli was knocked out in order to reduce the accumulation of acetic acid during tryptophan fermentation. The acetic acid content variation was investigated under various glucose levels. It showed that under high glucose content, the tdcD deletion strain YPTRP01 had a higher growth rate and exhibited increased tryptophan production in late exponential phase. The tryptophan yield was increased by 54.88% to 48.85g/L and the acetic acid level was reduced by 62.8% to 2.79g/L. In conclusion, the tdcD mutant showed more flexibility in the control of glucose contents during the tryptophan fermentation.