Lysine succinylation (Ksuc) is a novel protein post-translational modification (PTM) wherein a succinyl group modifies a lysine residue. Ksuc leads to significant chemical and structural changes to the modified protein. Recent studies have shown that Ksuc might play an important role in organism physiology and some pathophysiological processes, such as tumorigenesis and metabolic diseases. To provide an understanding of the molecular mechanism and functions of Ksuc in different organisms, we reviewed the current literature about Ksuc, mainly summarizing the research advances in eukaryotes and prokaryotes based on both traditional study methods and site prediction tools. We also discussed inhibitors or activators associated with Ksuc that may contribute to proteomic studies and could be useful in future clinical practice. A deeper understanding of Ksuc may shed new light on life science at the protein level and could lead to novel therapeutic strategies for various diseases.