Arbuscular mycorrhizal fungi (AMF) inoculation has important regulatory influence on plant growth and nitrogen uptake, which are intimately associated with soft wheat yield and quality. The study aimed to investigate the impact of AMF inoculation on soft wheat grain yield, protein compositions, protein quality, secondary structure of gluten, dough properties and biscuits baking quality. In this study, AMF inoculation significantly increased grain yield. AMF inoculation decreased grain protein content by 5.1%, glutenin content by 8.5%, GMP by 14.3%, and HMW-GS by 8.8% compared with CK. Conversely, AMF inoculation increased the ratio of gliadin/glutenin. Further analysis showed AMF treatment had a looser gluten network than CK, significantly reduced β-sheet and β-turn content, while α-helix content increased. The nutritional index of grain protein was slightly reduced under AMF inoculation, yet the biological value was improved. Reduced wet gluten content, gluten performance index and dough development time under AMF treatment through regulation of soft wheat protein composition and gluten protein structure. Therefore, biscuits in AMF treatment showed reduced thickness, hardness, significantly greater spread ratio and sensory score compared with CK. These indicate that AMF application would be a beneficial agronomic practice to synergistically optimize yield and grain quality in soft wheat.