Acetylation is a posttranslational modification conserved in all domains of life that is carried out by -acetyltransferases. While acetylation can occur on -amino groups, this review will focus on -acetylation of lysyl residues and how the posttranslational modification changes the cellular physiology of bacteria. Up until the late 1990s, acetylation was studied in eukaryotes in the context of chromatin maintenance and gene expression. At present, bacterial protein acetylation plays a prominent role in central and secondary metabolism, virulence, transcription, and translation. Given the diversity of niches in the microbial world, it is not surprising that the targets of bacterial protein acetyltransferases are very diverse, making their biochemical characterization challenging. The paradigm for acetylation in bacteria involves the acetylation of acetyl-CoA synthetase, whose activity must be tightly regulated to maintain energy charge homeostasis. While this paradigm has provided much mechanistic detail for acetylation and deacetylation, in this review we discuss advances in the field that are changing our understanding of the physiological role of protein acetylation in bacteria.