Cloned (Comment) | Organism |
---|---|
gene gapA, recombinant expression of His-tagged enzyme in Escherichia coli strain BL21(DE3) | Escherichia coli |
Crystallization (Comment) | Organism |
---|---|
purified recombinant enzyme GAPDH complexed with trehalose, hanging drop vapour diffusion method, mixing of 20 mg/ml protein in 20 mM Tris, 130 mM NaCl pH 7.5, with reservoir solution containing 2.8 M ammonium sulfate, 0.1 M MES, pH 5.5-6.5, 4°C, two to three weeks, GAPDH crystals are treated with a cryoprotectant consisting of 15% v/v trehalose at -173°C, X-ray diffraction structure determination and analysis at 2.1 A resolution, molecular replacement using the Escherichia coli GAPDH structure (PDB ID 1s7c) as a starting model | Escherichia coli |
Inhibitors | Comment | Organism | Structure |
---|---|---|---|
trehalose | might be an inhibitor, trehalose induces a conformational change in ecGAPDH in the current structure. The rotation of GAPDH also induced a conformational change in its active site. This suggests that the binding of trehalose to GAPDH induced a conformational change in its active site to prevent the binding of NAD+, although the NAD+- and trehalose-binding sites differ from one another | Escherichia coli |
Natural Substrates | Organism | Comment (Nat. Sub.) | Natural Products | Comment (Nat. Pro.) | Rev. | Reac. |
---|---|---|---|---|---|---|
D-glyceraldehyde 3-phosphate + phosphate + NAD+ | Escherichia coli | - |
3-phospho-D-glyceroyl phosphate + NADH + H+ | - |
? |
Organism | UniProt | Comment | Textmining |
---|---|---|---|
Escherichia coli | P0A9B2 | - |
- |
Purification (Comment) | Organism |
---|---|
recombinant His-tagged enzyme from Escherichia coli strain BL21(DE3) by nickel affinity chromatography, dialysis, and ultrafiltration | Escherichia coli |
Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
---|---|---|---|---|---|---|
D-glyceraldehyde 3-phosphate + phosphate + NAD+ | - |
Escherichia coli | 3-phospho-D-glyceroyl phosphate + NADH + H+ | - |
? |
Synonyms | Comment | Organism |
---|---|---|
EcGAPDH | - |
Escherichia coli |
GAPDH | - |
Escherichia coli |
glyceraldehyde-3-phosphate dehydrogenase | - |
Escherichia coli |
Temperature Stability Minimum [°C] | Temperature Stability Maximum [°C] | Comment | Organism |
---|---|---|---|
22 | - |
purified recombinant ecGAPDH is stable at room temperature | Escherichia coli |
Cofactor | Comment | Organism | Structure |
---|---|---|---|
NAD+ | - |
Escherichia coli |
General Information | Comment | Organism |
---|---|---|
additional information | the S-loop of GAPDH is required for interaction of the enzyme with its cofactor and with other proteins. NAD+-bound GAPDH S-loop fixation occurs by the formation of a complex with the coenzyme NAD+. The structure of trehalose-bound ecGAPDH is compared with the structures of both NAD+-free and NAD+-bound ecGAPDH. At the S-loop, the bound trehalose in the GAPDH structure induces a 2.4° rotation compared with the NAD+-free ecGAPDH structure and a 3.1° rotation compared with the NAD+-bound ecGAPDH structure | Escherichia coli |
physiological function | glyceraldehyde-3-phosphate dehydrogenase (GAPDH) catalyses the conversion of glyceraldehyde 3-phosphate to 1,3-bisphosphoglycerate using NAD+ as a cofactor. It is a moonlighting enzyme playing multiple roles in the regulation of mRNA stability, intracellular membrane trafficking, iron uptake and transport, DNA replication and repair, and nuclear RNA transport | Escherichia coli |