The glycerol-3-phosphate shuttle is a mechanism used in skeletal muscle and the brain[1] that regenerates NAD+ from NADH, a by-product of glycolysis. NADH is a reducing equivalent that stores electrons generated in the cytoplasm during glycolysis. NADH must be transported into the mitochondria to enter the oxidative phosphorylation pathway. However, the inner mitochondrial membrane is impermeable to NADH and only contains a transport system for NAD+. Depending on the type of tissue either the glycerol-3-phosphate shuttle pathway or the malate–aspartate shuttle pathway is used to transport electrons from cytoplasmic NADH into the mitochondria.[2]
The shuttle consists of two proteins acting in sequence. Cytoplasmic glycerol-3-phosphate dehydrogenase (cGPD) transfers an electron pair from NADH to dihydroxyacetone phosphate (DHAP), forming glycerol-3-phosphate (G3P) and regenerating the NAD+ needed to generate energy via glycolysis.[3] Mitochondrial glycerol-3-phosphate dehydrogenase (mGPD) then catalyzes the oxidation of G3P by FAD, regenerating DHAP in the cytosol and forming FADH2 in the mitochondrial matrix.[4] In mammals, its activity in transporting reducing equivalents across the mitochondrial membrane is secondary to the malate–aspartate shuttle.
Glycerol-3-phosphate is converted back to dihydroxyacetone phosphate by an inner membrane-bound mitochondrial glycerol-3-phosphate dehydrogenase 2 (GPD2 or mGPD), this time reducing one molecule of enzyme-bound flavin adenine dinucleotide (FAD) to FADH2. FADH2 then reduces coenzyme Q (ubiquinone to ubiquinol) whose electrons enter into oxidative phosphorylation.[12] This reaction is irreversible.[13] These electrons bypass Complex I of the electron transport chain, making the glycerol-3-phosphate shuttle less energetically efficient compared to oxidation of NADH by Complex I.[14]