Lysine acetylation exists extensively in prokaryotes, and plays a vital role in function adjustment. Recent progresses in the identification of prokaryote acetylation substrates and sites provide a great opportunity to explore the difference of substrate site specificity between prokaryotic and eukaryotic acetylation. Motif analysis suggests that prokaryotic and eukaryotic acetylation sites have distinct location-specific difference, and it is necessary to develop a prokaryote-specific acetylation sites prediction tool.