The present review addresses recent advances in research into a family of bifunctional enzymes that are responsible for the twofold task of synthesizing and hydrolyzing fructose-2,6-bisphosphate (Fru-2,6-P2), which in turn regulates the rate of glycolysis in most cells. The structure of the synthetic kinase, conjoined at its carboxyl-terminus to the phosphatase, is very highly conserved throughout evolution and differentiation, with isotypic expression arising from highly variable amino-terminal and carboxyl-terminal regulatory domains. These domains, which frequently contain protein-kinase-catalyzed phosphorylation motifs, are responsible for the widely divergent kinetics observed in various tissues and species, and for the hormonal modulation that alters intracellular levels of Fru-2,6-P2. The present review discusses recent advances in relating structure to function, and the identification of new pathways of transcriptional regulation of this important family of regulatory enzymes.