Consumers with gluten-related disorders require gluten-free (GF) foods to avoid an immune response. Alternative to the use of non-gluten containing grains to prepare GF bread, the gluten reactivity has been greatly reduced using a proline specific cleavage enzyme, however, the gluten functionality was lost. The aim of this study was to evaluate the effect of adding an amaranth flour blend (AFB) to enzymatically modified wheat-flour proteins on dough functionality and to evaluate the immunoreactivity and acceptability of the prepared bread. First, wheat flour (20% w/v, substrate) was hydrolyzed using 8.4 U mg-1 protein Aspergillus niger prolyl-endopeptidase (AnPEP) for 8 h at 40 °C under constant agitation. Four types of breads were prepared with the same formulation except for the type of flour (14% w.b.): wheat flour (WF), WF-AFB unmodified not incubated, WF-AFB unmodified incubated and WF-AFB modified. The protein composition and free thiols were analyzed before and after amaranth addition, and the flour and bread proteins were run using SDS-PAGE and immune-detected in blots with IgA from celiac disease patients. The immunoreactive gluten content, specific volume and bread acceptability were evaluated. The polymeric proteins and free thiol groups of WF decreased after AnPEP treatment. The electrophoretic patterns of the modified flour and bread proteins were different and the IgA-immunodetection in blots was highly reduced, particularly for the higher molecular weight subunits. The addition of AFB to the modified wheat flour prepared using AnPEP improved the dough functionality by increasing the thiol groups and allowed the preparation of a sensorially acceptable bread with only 60 mg kg-1 immunoreactive gluten.