Glyceraldehyde 3-phosphate dehydrogenase

Glyceraldehyde 3-phosphate dehydrogenase (abbreviated as GAPDH or less commonly as G3PDH) (EC 1.2.1.12) is an enzyme of ~37kDa that catalyzes the sixth step of glycolysis and thus serves to break down glucose for energy and carbon molecules. In addition to this long established metabolic function, GAPDH has recently been implicated in several non-metabolic processes, including transcription activation, initiation of apoptosis, ER to Golgi vesicle shuttling, and fast axonal, or axoplasmic transport. In sperm, a testis-specific isoenzyme GAPDHS is expressed. Under normal cellular conditions, cytoplasmic GAPDH exists primarily as a tetramer. This form is composed of four identical 37-kDa subunits containing a single catalytic thiol group each and critical to the enzyme's catalytic function. Nuclear GAPDH has increased isoelectric point (pI) of pH 8.3–8.7. Of note, the cysteine residue C152 in the enzyme's active site is required for the induction of apoptosis by oxidative stress. Notably, post-translational modifications of cytoplasmic GAPDH contribute to its functions outside of glycolysis. GAPDH is encoded by a single gene that produces a single mRNA transcript with no known splice variants, though an isoform does exist as a separate gene that is expressed only in spermatozoa. Compound C00118 at KEGG Pathway Database. Enzyme 1.2.1.12 at KEGG Pathway Database. Reaction R01063 at KEGG Pathway Database. Compound C00236 at KEGG Pathway Database. The first reaction is the oxidation of glyceraldehyde 3-phosphate (G3P) at the position-1 (in the diagram it is shown as the 4th carbon from glycolysis), in which an aldehyde is converted into a carboxylic acid (ΔG°'=-50 kJ/mol (−12kcal/mol)) and NAD+ is simultaneously reduced endergonically to NADH. The energy released by this highly exergonic oxidation reaction drives the endergonic second reaction (ΔG°'=+50 kJ/mol (+12kcal/mol)), in which a molecule of inorganic phosphate is transferred to the GAP intermediate to form a product with high phosphoryl-transfer potential: 1,3-bisphosphoglycerate (1,3-BPG). This is an example of phosphorylation coupled to oxidation, and the overall reaction is somewhat endergonic (ΔG°'=+6.3 kJ/mol (+1.5)). Energy coupling here is made possible by GAPDH. GAPDH uses covalent catalysis and general base catalysis to decrease the very large and positive activation energy of the second step of this reaction. First, a cysteine residue in the active site of GAPDH attacks the carbonyl group of GAP, creating a hemithioacetal intermediate (covalent catalysis). Next, an adjacent, tightly bound molecule of NAD+ accepts a hydride ion from GAP, forming NADH while GAP is simultaneously oxidized to a thioester in a concerted series of steps. This thioester species is much higher in energy than the carboxylic acid species that would result in the absence of GAPDH (the carboxylic acid species is so low in energy that the energy barrier for the second step of the reaction (phosphorylation) would be too high, and the reaction, therefore, too slow and equilibrium too unfavorable for a living organism). Donation of the hydride ion by the hemithioacetal is facilitated by its deprotonation by a histidine residue in the enzyme's active site (general base catalysis). Deprotonation encourages the reformation of the carbonyl group in the thioester intermediate and ejection of the hydride ion. NADH leaves the active site and is replaced by another molecule of NAD+, the positive charge of which stabilizes the negatively charged carbonyl oxygen in the transition state of the next and ultimate step. Finally, a molecule of inorganic phosphate attacks the thioester and forms a tetrahedral intermediate, which then collapses to release 1,3-bisphosphoglycerate, and the thiol group of the enzyme's cysteine residue.