Using the inhibitors chloramphenicol and cycloheximide, the relative importance of mitochondrial vs. cytosolic protein synthesis to brewing yeast performance and final beer quality was assessed. Cytosolic protein synthesis was much more critical, but mitochondrial protein synthesis did affect the overall performance of a commercial lager strain of Saccharomyces cerevisiae in high-gravity (16° Plato) brewers' wort. Previous research had indicated a critical role for mitochondrial energy in the promotion of adequate fermentation despite the fact that brewing yeasts reportedly obtained all of their energy from fermentative growth. This research indicated that mitochondrial protein synthesis was involved in yeast growth, specific carbohydrate uptake rates, free amino nitrogen utilization, ethanol production, and lipid and glycogen biosynthesis. This was true for yeast in two different physiological conditions used for inoculation. Artificially aged yeast showed a greater susceptibility to the inhibitors than did brewery-fresh yeast, presumably due to permeability differences and/or a lack of ability for the compromised cells to counteract the inhibitor damage. The blockage of mitochondrial protein synthesis did not alter the yeast performance as radically as did inhibiting the energy supply to these organelles but did inhibit the commercial yeast strain's ability to ferment optimally. It is suggested that mitochondrial protein synthesis occurs during the brewing fermentation and contributes to the efficiency and end quality of the product and the yeast harvested from the fermentation for reuse. However, protein synthesis in the mitochondria was not the major contributor to previous observations, indicating a critical role for mitochondria in brewing yeast performance.