Technical Abstract: In a dry grind ethanol plant, approximately 0.84 kg of dried distillers’ grains with solubles (DDGS) is produced per liter of ethanol. The distillers’ grains contain the unhydrolyzed and unprocessed cellulosic fraction of corn kernels, which could be further converted to ethanol or other valuable bioproducts by applying cellulose conversion technology. Its compositional variability is one of the factors that could affect the overall process design and economics. In this study, we present compositional variability of distillers’ grains collected from four different dry grind ethanol plants and its effect on enzymatic digestibility and fermentability. We then selected two sources of distillers' grains based on their distinctive compositional difference. These were pretreated by either controlled pH liquid hot water (LHW) or ammonia fiber expansion (AFEX) and subjected to enzymatic hydrolysis and fermentation. Fermentation of the pretreated distillers’ grains using either industrial yeast or genetically engineered glucose and xylose co-fermenting yeast, yielded 70–80% of theoretical maximum ethanol concentration, which varied depending on the batch of distillers’ grains used. Results show that cellulose conversion and ethanol fermentation yields are affected by the compositions of distillers’ grains. Distillers’ grains with a high extractives content exhibit a lower enzymatic digestibility but a higher fermentability.