Due to the desire for nutritionally dense foods, there is interest in whole grain and sourdough bread products. This research study was carried out to evaluate the effect of different starter cultures and stone mill settings on the nutritional attributes of sourdough bread. Using a stone mill, a blend of three hard red spring wheat varieties was ground into whole-grain flour. Three different gap settings and two different rotation speed values were used to produce flour samples with six different treatments, and those six flour samples were used to make sourdough bread with two different starter cultures. The experimental design was a randomized complete block design with a factorial arrangement. Significant differences (p < 0.05) were observed in dietary fiber content, arabinoxylan content, total starch content, and protein digestibility between sourdough bread made with rye and wheat starter cultures. Higher protein digestibility values were observed for most of the sourdough samples with the rye starter culture compared to the wheat starter culture. In addition, the protein digestibility showed a strong negative correlation (r = −0.85) with the coarse particles and a positive correlation (r = 0.70) with the fine particle size fraction. These results demonstrated that selected starter cultures and the raw material with the desired particle size could improve sourdough bread's nutritional attributes and potential application in the baking industry.