AbstractProtein kinases have become the second most important group of drug targets for the pharmaceutical industry next to G‐protein‐coupled receptors. Thus, over the past decade, a significant number of small molecules have been generated for protein kinase drug optimization programs. The vast majority of kinase inhibitors target the ATP binding site of the enzyme; however, the poor protein kinase selectivity of ATP‐competitive protein kinase inhibitors (PKIs) limits their use for treating chronic diseases. In contrast, for inhibitors of bacterial signal transduction systems targeting bacterial kinase(s), there are no such selectivity requirements as long as the inhibitor does not act on any human kinases at the effective concentrations for killing bacteria in vivo. Protein phosphorylation in bacteria is performed by two‐component signal transduction systems (2CSTSs) and eukaryotic‐like serine/threonine kinases or bacterial tyrosine kinases. Recently, a large number of studies of protein kinases essential for sustaining bacterial growth and kinases required for virulence have been reported. Thus, bacterial protein kinases offer considerable potential as new drug targets. To identify bacterial PKIs, large chemical libraries of ATP‐competitive inhibitors developed for eukaryotic protein kinases are an invaluable asset. This manuscript reviews progress on the development of prokaryotic protein kinase inhibitors. Drug Dev Res 2010. © 2010 Wiley‐Liss, Inc.