Survival in extreme environments requires drastic changes to animals’ physiology and biochemistry, requiring many animals to decrease their metabolic rate. Enzymes are the mediators of metabolic functions and their regulation is critical to surviving these conditions. This thesis explores the properties of a key enzyme of glycolysis, pyruvate kinase (PK), in the freeze tolerant wood frog (Rana sylvatica), the anoxia tolerant snail (Littorina littorea) and the hibernating ground squirrel (Urocitellus richardsonii).The studies showed that temperature, allosteric effectors and other local environmental conditions play a role in regulating pyruvate kinase in response to freezing, anoxia and during hibernation. Furthermore, reversible protein phosphorylation appears to be a common regulatory mechanism of PK by reducing the activity of the enzyme in all three animals. Together, these studies show that PK is a common regulatory target of animals surviving in extreme conditions and that multiple regulatory mechanisms, especially protein phosphorylation, contribute to its regulation.