Protein Nε-acetylation, a post-translational modification widespread in eukaryotes and prokaryotes, has been intensively explored due to its crucial roles in multitudinous physiologic events including transcriptional regulation, metabolic regulation, etc. The particular hotspot is the relationship between acetylation and metabolic regulation. Protein acetylation major types and functions thereof, prokaryotic acetyltransferase, deacetylases, and acetylation sites of enzymes related to glycometabolism, together with the cross-talk between acetylation and other modification, such as the phosphorylation were summarized, with emphases on those from Mycobacteria. J. Cell. Biochem. 113: 3601–3609, 2012. © 2012 Wiley Periodicals, Inc.