Abstract: γ-aminobutyric acid (GABA) is a major inhibitory neurotransmitter in the central nervous system and plays important roles in alleviating insomnia, depression and nervous disorders. In order to increase the content of GABA content in steamed bread and improve the flavor and texture, strains of Levilactobacillus sp. LB-2 (with high GABA production ability), Saccharomyces cerevisiae Y5 and Acetobacter sp. A3 was selected to make steamed bread by single strain and mixed-strain fermentation, and the effects of mixed-strain fermentation on the specific volume, color, textural characteristics, and other edible qualities of the steamed bread enriched in GABA were also investigated. The results showed that when the mixed fermentation of Saccharomyces cerevisiae Y5, Levilactobacillus sp. LB-2 and Acetobacter sp. A3 were in the dough starter, the GABA content (6.16 mg/g) of the steamed was the highest in YLA treatment, which was about 61.3% higher than that of the steamed bread (3.82 mg/g) fermented by single yeast. But the addition of corn flour and scalding process were not conducive to the production of GABA content in the steamed bread. When the cell ratio of Saccharomyces cerevisiae Y5, Levilactobacillus sp. LB-2 and Acetobacter sp. A3 were 1: 5: 1 in the dough starter, the specific volume, height-to-diameter ratio and L^* value of the steamed bread were significantly higher than the rest of the treatment groups (P<0.05), while the hardness was significantly lower (P<0.05), and the hardness of the steamed bread with scalding process was the lowest, which indicated that the scalding process contributed to the improvement of the quality of the steamed bread. The results of this study could lay a foundation for the development of GABA-rich steamed bread or other kinds of food products made by fermented dough.