Literature summary for 1.17.1.1 extracted from

Gonzalez-Porque, P.; Strominger, J.L.
Enzymatic synthesis of cytidine diphosphate 3,6-dideoxyhexoses. VI. Purification to homogeneity and some properties of cytidine diphosphate-D-glucose oxidoreductase, enzyme E1 and enzyme E3 (1972), J. Biol. Chem., 247, 6748-6756.

Inhibitors

Inhibitors	Comment	Organism	Structure
5,5'-dithiobis(2-nitrobenzoate)	-	Yersinia pseudotuberculosis
N-ethylmaleimide	enzyme E1: dithiothreitol in excess protects against inhibition	Yersinia pseudotuberculosis

Molecular Weight [Da]

Molecular Weight [Da]	Molecular Weight Maximum [Da]	Comment	Organism
35000	45000	enzyme E3, thin-layer chromatography on Sephadex G-100	Yersinia pseudotuberculosis
40000	-	enzyme E3: 1 * 40000, SDS-PAGE, enzyme E1: 1 * 61000, SDS-PAGE, 2 proteins E1 and E3 are involved but no partial reaction has been observed in the presence of either alone	Yersinia pseudotuberculosis
50000	70000	enzyme E1, thin-layer chromatography on Sephadex G-100	Yersinia pseudotuberculosis
61000	-	enzyme E3: 1 * 40000, SDS-PAGE, enzyme E1: 1 * 61000, SDS-PAGE, 2 proteins E1 and E3 are involved but no partial reaction has been observed in the presence of either alone	Yersinia pseudotuberculosis

Organism

Organism	UniProt	Comment	Textmining
Yersinia pseudotuberculosis	-	25 VO	-
Yersinia pseudotuberculosis 25VO	-	25 VO	-

Purification (Commentary)

Purification (Comment)	Organism
the first three steps are common to the purification of enzyme E1, enzyme E3 and cofactor, their separation can be accomplished after step 4, step 1: preparation of the crude extract, step 2: streptomycin sulfate precipitation, step 3: ammonium sulfate precipitation and dialysis, step 4: DEAE-cellulose chromatography, step 5: purification of enzyme E1, gel filtration on Sephadex G-100, step 6: second DEAE-cellulose chromatography, step 7: preparative polyacrylamide gel electrophoresis, step 5': purification of enzyme E3, gel filtration on Sephadex G-100, step 6': second DEAE-cellulose chromatography, step 7': third DEAE-cellulose chromatography	Yersinia pseudotuberculosis

Purification (Comment)

Organism

the first three steps are common to the purification of enzyme E1, enzyme E3 and cofactor, their separation can be accomplished after step 4, step 1: preparation of the crude extract, step 2: streptomycin sulfate precipitation, step 3: ammonium sulfate precipitation and dialysis, step 4: DEAE-cellulose chromatography, step 5: purification of enzyme E1, gel filtration on Sephadex G-100, step 6: second DEAE-cellulose chromatography, step 7: preparative polyacrylamide gel electrophoresis, step 5': purification of enzyme E3, gel filtration on Sephadex G-100, step 6': second DEAE-cellulose chromatography, step 7': third DEAE-cellulose chromatography

Yersinia pseudotuberculosis

Reaction

Reaction	Comment	Organism	Reaction ID
CDP-4-dehydro-3,6-dideoxy-D-glucose + NAD(P)+ + H2O = CDP-4-dehydro-6-deoxy-D-glucose + NAD(P)H + H+	mechanism	Yersinia pseudotuberculosis	a
CDP-4-dehydro-3,6-dideoxy-D-glucose + NAD(P)+ + H2O = CDP-4-dehydro-6-deoxy-D-glucose + NAD(P)H + H+	E1 binds the substrate pyridoxamine 5'-phosphate essential for binding, E3 possesses NADH oxidase activity, may be the reductase	Yersinia pseudotuberculosis	a
CDP-4-dehydro-3,6-dideoxy-D-glucose + NAD(P)+ + H2O = CDP-4-dehydro-6-deoxy-D-glucose + NAD(P)H + H+	role of the enzymes E1 and E3, as well as role of the cofactor. Multicomponent system consisting of substrate, NADH or NADPH, enzyme E1: recognition unit of the system by binding to the substrate through the amino group of the coenzyme, enzyme E3: acts as the dehydrogenase of the system, and cofactor: pyridoxamine 5'-phosphate	Yersinia pseudotuberculosis	a

Specific Activity [micromol/min/mg]

Specific Activity Minimum [µmol/min/mg]	Specific Activity Maximum [µmol/min/mg]	Comment	Organism
additional information	-	-	Yersinia pseudotuberculosis

Storage Stability

Storage Stability	Organism
lyophilized powder: enzyme E3 not stable	Yersinia pseudotuberculosis
lyophilized powder: for months, enzyme E1	Yersinia pseudotuberculosis

Subunits

Subunits	Comment	Organism
monomer	enzyme E3: 1 * 40000, SDS-PAGE, enzyme E1: 1 * 61000, SDS-PAGE, 2 proteins E1 and E3 are involved but no partial reaction has been observed in the presence of either alone	Yersinia pseudotuberculosis

Temperature Optimum [°C]

Temperature Optimum [°C]	Temperature Optimum Maximum [°C]	Comment	Organism
25	-	assay at	Yersinia pseudotuberculosis

pH Optimum

pH Optimum Minimum	pH Optimum Maximum	Comment	Organism
7.5	-	assay at	Yersinia pseudotuberculosis

Cofactor

Cofactor	Comment	Organism
additional information	4.3 SH per mol of enzyme E1 in the presence of SDS, calculated using a molecular weight of 61000 Da	Yersinia pseudotuberculosis
additional information	0.37 SH groups per molecule of enzyme E3, in the absence of SDS and 1 SH group per molecule of enzyme E3 in the presence of SDS, the single SH group in enzyme E3 is essential for activity	Yersinia pseudotuberculosis
pyridoxamine 5'-phosphate	bound to enzyme E1 through an ionic interaction with a positive charge on the surface of the enzyme, the cofactor is needed for the binding of the substrate to the enzyme	Yersinia pseudotuberculosis