Mycobacterium tuberculosis parasitizes resting macrophages yet is killed by activated macrophages through both oxidative and nonoxidative mechanisms. Nonoxidative mechanisms are linked to the maturation of the bacteria-containing phagosome into an acidified, hydrolytically active compartment. We describe here a mechanism for killing Mycobacteria in the lysosomal compartment through the activity of peptides generated by the hydrolysis of ubiquitin. The induction of autophagy in infected macrophages enhanced the delivery of ubiquitin conjugates to the lysosome and increased the bactericidal capacity of the lysosomal soluble fraction. The accumulation of ubiquitinated proteins in the autophagolysosome provides one possible mechanism behind the antimicrobial activities observed for a range of pathogens in autophagous host cells.