Which are the three irreversible steps of glycolysis and their enzymes?

• A. Hexokinase/glucokinase; Phosphofructokinase-1; Pyruvate kinase
• B. Hexokinase; Pyruvate dehydrogenase; Citrate synthase
• C. Glucokinase; Aldolase; Enolase
• D. Phosphoglycerate mutase; Glyceraldehyde-3-phosphate dehydrogenase; 3-phosphoglycerate kinase

Answer: (A)

Three irreversible steps in glycolysis are those that commit glucose to breakdown because they release a large amount of free energy and are tightly regulated. The first is the phosphorylation of glucose to glucose-6-phosphate by hexokinase (or glucokinase in liver and pancreatic cells), which traps glucose in the cell and uses one ATP. The second is the committed step, the conversion of fructose-6-phosphate to fructose-1,6-bisphosphate by phosphofructokinase-1, a highly regulated, highly exergonic reaction that gates glycolysis. The third is the formation of pyruvate from phosphoenolpyruvate by pyruvate kinase, producing ATP in a substrate-level phosphorylation step and driving the pathway forward. These three steps are effectively irreversible under cellular conditions, while other glycolytic steps either operate near equilibrium or feed into other pathways; for example, enzymes like pyruvate dehydrogenase and citrate synthase connect glycolysis to the TCA cycle, and steps such as aldolase, enolase, glyceraldehyde-3-phosphate dehydrogenase, and 3-phosphoglycerate kinase can be reversible depending on conditions.