Fermentation is largely a physiological balancing act.  During the process one portion of a substrate is oxidized while another portion is reduced.  Moreover, fermentation is the result of concerted activities among many different enzymes rather than the product of one or two.  Overall metabolic flux is rarely determined by the activity of any one enzyme in a pathway, and activities of multiple enzymes often interact.  It should not be surprising, therefore, that the engineering of one or two or even a few enzymes in a metabolic pathway does not approach an optimum.  Our laboratory has developed techniques for balancing the activities of multiple enzymes in the pathway for yeast xylose fermentations, for discovering novel activities that contribute to improved fermentation rates and for predicting steps that could be rate limiting.  In engineered Saccharomyces cerevisiae and Pichia stipitis, the balance between oxidative and reductive activities can determine the flux of xylose to either xylitol or ethanol.  The overall, flux, however, is generally determined by the activities of “gateway” enzymes such as kinases or transporters. The required balance of activities can be predicted from global expression analysis of cells with various genetic backgrounds cultivated under different growth conditions.  This presentation will review the transcriptional and physiological responses of native and engineered xylose and cellobiose fermenting yeasts during ethanol production.