The second step of gluconeogenesis occurs in the mitochondria with the irreversible conversion of oxaloacetate to phosphoenolpyruvate by PEP carboxykinase.
Gluconeogenesis uses many of the same enzymes as glycolysis, but in the opposite direction since many of these reactions are reversible. However, three steps require bypassing since they are performed by irreversible enzymatic reactions. The first irreversible step requires two reactions, the conversion of pyruvate to oxaloacetate by pyruvate carboxylase in the mitochondria followed by the conversion of oxaloacetate to phosphoenolpyruvate by PEP carboxykinase in the same location. The second irreversible step is the conversion of fructose-1,6-biphosphate to fructose-6-phosphate by fructose-1,6-bisphosphatase in the cytosol. The final step is the dephosphorylation of glucose-6-phosphate to glucose by glucose-6-phosphatase in the endoplasmic reticulum.
Berg et al. describe the regulation of glycolysis and gluconeogenesis at various steps. The interconversion of fructose-6-phosphate to fructose-1,6-bisphosphate (the second step mentioned above) is regulated by the levels of AMP, ATP, and citrate. A high level of ATP and citrate, signals that the cell has sufficient energy, switches gluconeogenesis on and turns off glycolysis by activating fructose-1,6-biphosphatase and inhibiting phosphofructokinase, respectively. The converse occurs when there is a high level of AMP, a signal that the cell has a low level of energy, turning off gluconeogenesis and activates glycolysis. (See illustration A for more details)
Berg et al. also describe a second mechanism that tightly regulates the same enzymes, phosphofructokinase and fructose-1,6-bisphosphatase, through the action of fructose-2,6-bisphosphate (F-2,6-BP). The amount of F-2,6-BP varies depending on the fed (high F-2,6-BP) and fasting (low F-2,6-BP) state of the cell. F-2,6-BP stimulates phosphofructokinase and inhibits fructose-1,6-bisphosphatase, switching on glycolysis during the fed state. During starvation, low F-2,6-BP switches on gluconeogenesis and inhibits glycolysis.
Illustration A demonstrates the factors that regulate two of the irreversible steps in glycolysis/gluconeogenesis.
Answer 1: The irreversible conversion of glucose-6-phosphate to glucose by glucose-6-phosphatase during gluconeogenesis occurs in the endoplasmic reticulum.
Answer 2: The irreversible conversion of fructose-1,6-biphosphate to fructose-6-phosphate by fructose-1,6-bisphosphatase during gluconeogenesis occurs in the cytosol.
Answer 4: The irreversible conversion of phosphoenolpyruvate to pyruvate by pyruvate kinase during glycolysis occurs in the cytosol.
Answer 5: The irreversible conversion of glucose-6-phosphate to 6-phosphogluconolactone by glucose-6-phosphate dehydrogenase during the hexose monophosphate shunt occurs in the cytosol.
Berg JM, Tymoczko JL, Stryer L. Biochemistry. 5th edition. New York: W H Freeman; 2002. Section 16.4, Gluconeogenesis and Glycolysis Are Reciprocally Regulated. Available from: http://www.ncbi.nlm.nih.gov/books/NBK22423/