Literature summary for 1.2.1.12 extracted from

Rodriguez-Hernandez, A.; Romo-Arevalo, E.; Rodriguez-Romero, A.
A novel substrate-binding site in the X-ray structure of an oxidized E. coli glyceraldehyde 3-phosphate dehydrogenase elucidated by single-wavelength anomalous dispersion (2019), Crystals, 9, 622 .

No PubMed abstract available

Application

Application	Comment	Organism
analysis	GAPDH is a multi-functional protein that is used as a control marker for basal function, it is known to undergo cysteine oxidation under different types of cellular stress	Escherichia coli

Cloned(Commentary)

Cloned (Comment)	Organism
recombinant expression of His-tagged enzyme in Escherichia coli strain BL21(DE3)	Escherichia coli

Crystallization (Commentary)

Crystallization (Comment)	Organism
native, non-modified and selenium-modified enzymes, 5.5 mg/ml protein in 10 mM HEPES, pH 7.5, and 1.43 M sodium citrate, 18°C, 3 days, method optimization, crystals from the selenium modified enzyme are soaked for 10 min in 5 mM NAD+ dissolved in mother liquor with or without 25% trehalose, X-ray diffraction structure determination and analysis at 1.64-2.14 A resolution using single-wavelength anomalous dispersion (SAD) phasing with a selenium-modified enzyme, molecular replacement	Escherichia coli

Crystallization (Comment)

Organism

native, non-modified and selenium-modified enzymes, 5.5 mg/ml protein in 10 mM HEPES, pH 7.5, and 1.43 M sodium citrate, 18°C, 3 days, method optimization, crystals from the selenium modified enzyme are soaked for 10 min in 5 mM NAD+ dissolved in mother liquor with or without 25% trehalose, X-ray diffraction structure determination and analysis at 1.64-2.14 A resolution using single-wavelength anomalous dispersion (SAD) phasing with a selenium-modified enzyme, molecular replacement

Escherichia coli

Protein Variants

Protein Variants	Comment	Organism
additional information	generation of an engineered synthetic Escherichia coli codon optimized sequence of a human gene that codifies for a 32.4 kDa protein for recombinant expression	Escherichia coli

Inhibitors

Inhibitors	Comment	Organism	Structure
H2O2	irreversible inhibition	Escherichia coli

Localization

Localization	Comment	Organism	GeneOntology No.	Textmining
cytoplasm	-	Escherichia coli	5737	-

Natural Substrates/ Products (Substrates)

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

Organism	UniProt	Comment	Textmining
Escherichia coli	P0A9B2	-	-

Purification (Commentary)

Purification (Comment)	Organism
recombinant His-tagged engineered enzyme from Escherichia coli strain BL21(DE3) by nickel affinity chromatography, gel filtration, and ultrafiltration	Escherichia coli

Substrates and Products (Substrate)

Substrates	Comment Substrates	Organism	Products	Comment (Products)	Rev.	Reac.
D-glyceraldehyde 3-phosphate + phosphate + NAD+	-	Escherichia coli	3-phospho-D-glyceroyl phosphate + NADH + H+	-	?
D-glyceraldehyde 3-phosphate + phosphate + NAD+	substrate binding structure analysis, overview	Escherichia coli	3-phospho-D-glyceroyl phosphate + NADH + H+	-	?

Subunits

Subunits	Comment	Organism
tetramer	4 * 32400, engineered recombinant enzyme, SDS-PAGE, 4 * 35500, non-modified wild-type enzyme, SDS-PAGE	Escherichia coli

Synonyms

Synonyms	Comment	Organism
EcGAPDH	-	Escherichia coli
GapA	-	Escherichia coli
GAPDH	-	Escherichia coli
glyceraldehyde 3-phosphate dehydrogenase	-	Escherichia coli

Cofactor

Cofactor	Comment	Organism	Structure
NAD+	binding site and binding structure, detailed overview. The NAD+ binding domain consist of six parallel and one anti-parallel beta-strands, surrounded by four alpha-helices, the typical structure of a Rossmann fold	Escherichia coli

General Information

General Information	Comment	Organism
malfunction	when the cell is exposed to high levels of H2O2, GAPDH is irreversibly inhibited presumably by the formation of sulphenic acid in the active site cysteine, becoming a switch that balances the equilibrium between the glycolytic cycle and the pentose phosphate metabolic pathway and promoting the formation of NADPH to combat ROS-produced cell stress	Escherichia coli
additional information	each GAPDH monomer contains a molecule of glyceraldehyde-3 phosphate in a non-previously identified site. The catalytic Cys149 is covalently attached to an about 300 Da molecule, possibly glutathione. This modification alters the conformation of an adjacent alpha-helix in the catalytic domain, right opposite to the NAD+ binding site. The conformation of the alpha-helix is stabilized after soaking the crystals with NAD+. Enzyme structure analysis, structure modeling, detailed overview	Escherichia coli
physiological function	apart from its glycolytic function, GAPDH displays a battery of moonlighting activities. Primary location of the tetrameric GAPDH is in the cytoplasm, where it conducts its canonical role in glycolysis	Escherichia coli