Abstract Solving the issue of the origin of life on Earth is impossible without understanding how the chemical, functional and regulatory principles underlying cellular metabolism arose, how cells acquired the properties that determine their evolution, and how biological systems function and develop. This review addresses functional versatility of glycolytic enzymes whose expression is considerably increased in some types of cells, for example, stem or malignant tumor cells. Almost all glycolytic enzymes share non-catalytic functions needed to maintain a high rate of cell proliferation, their active migration, and the formation of a stem-like phenotype. Glycolytic enzymes emerged very early in evolution. Since the genomes of ancient life forms contained a limited number of genes to encode all the multitude of required functions, glycolytic enzymes or the products of the reactions they catalyzed could be exploited as ancient regulators of inter- and intracellular communications. Subsequently, the multifunctionality of the main metabolic enzymes began to be used by tumor cells to ensure their survival and growth. In this review, we discuss some of the non-catalytic functions of glycolytic enzymes, as well as the possible evolutionary significance of acquiring such a multifunctionality.