Literature summary for 1.1.1.205 extracted from

Labesse, G.; Alexandre, T.; Gelin, M.; Haouz, A.; Munier-Lehmann, H.
Crystallographic studies of two variants of Pseudomonas aeruginosa IMPDH with impaired allosteric regulation (2015), Acta Crystallogr. Sect. D, 71, 1890-1899.

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Activating Compound

Activating Compound	Comment	Organism	Structure
MgATP2-	allosteric regulation of Pseudomonas aeruginosa IMPDH by MgATP2-. The the ATP-binding sites are located within the two CBS motifs. The compound acts as a positive effector increasing the catalytic activity of the wild-type enzyme, and also of the D199N variant, influencing both the maximal rate and the affinity for IMP, and showing a cooperativity effect for IMP, that is lost in mutant D199N	Pseudomonas aeruginosa

Cloned(Commentary)

Cloned (Comment)	Organism
gene guaB, sequence comparisons, recombinant expression of wild-type and mutant enzymes in Escherichia coli	Pseudomonas aeruginosa

Crystallization (Commentary)

Crystallization (Comment)	Organism
purified recombinant IMPDH enzyme mutant DELTACBS in the apo form and in complex with IMP, sitting drop vapour diffusion method, mixing of 0.0015 ml of 9.2 mg/ml protein in 20 mM potassium phosphate, pH 8.0, 25 mM KCl, with 0.0015 ml reservoir solution consisting of either 4.3 M sodium chloride, 100 mM HEPES, pH 7.5, or 10 mM sodium citrate, and 33% PEG 8000 for the apoenzyme and containing 1.26 M ammonium sulfate, 100 mM HEPES, pH 7.5, for the IMP-complexed enzyme, equilibration against 0.15 ml or reservoir solution. For the D199N variant in complex with IMP, the optimized crystals are obtained by mixing 0.0015 ml of 11.7 mg/ml protein in solution in 20 mM potassium phosphate, pH 8.0, 25 mM KCl, and 10 mM IMP, with 0.0015 ml of reservoir solution containing 10% w/v PEG 4000, 200 mM magnesium chloride, 100 mM MES pH 6.5, and equilibration against 0.15 ml or reservoir solution, 18°C, X-ray diffraction structure determination and analysis at 1.65-1.95 A resolution, molecular replacement using a previously solved structure of wild-type IMPDHpa, PDB ID 4dqw, as a template	Pseudomonas aeruginosa

Crystallization (Comment)

Organism

purified recombinant IMPDH enzyme mutant DELTACBS in the apo form and in complex with IMP, sitting drop vapour diffusion method, mixing of 0.0015 ml of 9.2 mg/ml protein in 20 mM potassium phosphate, pH 8.0, 25 mM KCl, with 0.0015 ml reservoir solution consisting of either 4.3 M sodium chloride, 100 mM HEPES, pH 7.5, or 10 mM sodium citrate, and 33% PEG 8000 for the apoenzyme and containing 1.26 M ammonium sulfate, 100 mM HEPES, pH 7.5, for the IMP-complexed enzyme, equilibration against 0.15 ml or reservoir solution. For the D199N variant in complex with IMP, the optimized crystals are obtained by mixing 0.0015 ml of 11.7 mg/ml protein in solution in 20 mM potassium phosphate, pH 8.0, 25 mM KCl, and 10 mM IMP, with 0.0015 ml of reservoir solution containing 10% w/v PEG 4000, 200 mM magnesium chloride, 100 mM MES pH 6.5, and equilibration against 0.15 ml or reservoir solution, 18°C, X-ray diffraction structure determination and analysis at 1.65-1.95 A resolution, molecular replacement using a previously solved structure of wild-type IMPDHpa, PDB ID 4dqw, as a template

Pseudomonas aeruginosa

Protein Variants

Protein Variants	Comment	Organism
D199N	site-directed mutagenesis, mutant structure analysis	Pseudomonas aeruginosa
additional information	construction of a mutant lacking the CBS domains, IMPDH mutant DELTACBS. The DELTACBS variant lacks the CBS motifs from Ala92 to Lys202 of the full-length enzyme. This variant is fully active and its kinetic parameters are similar to those of wild-type IMPDHpa in the presence of its positive effector Mg-ATP. The catalytic activity of the D199N variant is still sensitive to Mg-ATP, influencing both the maximal rate and the affinity for IMP, but the cooperativity effect for IMP is lost	Pseudomonas aeruginosa

KM Value [mM]

KM Value [mM]	KM Value Maximum [mM]	Substrate	Comment	Organism	Structure
additional information	-	additional information	Michaelis-Menten kinetics	Pseudomonas aeruginosa

Metals/Ions

Metals/Ions	Comment	Organism	Structure
MgATP2-	allosteric regulation of Pseudomonas aeruginosa IMPDH by MgATP2-	Pseudomonas aeruginosa

Natural Substrates/ Products (Substrates)

Natural Substrates	Organism	Comment (Nat. Sub.)	Natural Products	Comment (Nat. Pro.)	Rev.	Reac.
IMP + NAD+ + H2O	Pseudomonas aeruginosa	-	XMP + NADH + H+	-	?

Organism

Organism	UniProt	Comment	Textmining
Pseudomonas aeruginosa	Q9HXM5	-	-

Purification (Commentary)

Purification (Comment)	Organism
recombinant wild-type and mutant enzymes from Escherichia coli by affinity chromatography and gel filtration	Pseudomonas aeruginosa

Substrates and Products (Substrate)

Substrates	Comment Substrates	Organism	Products	Comment (Products)	Rev.	Reac.
IMP + NAD+ + H2O	-	Pseudomonas aeruginosa	XMP + NADH + H+	-	?

Subunits

Subunits	Comment	Organism
More	IMPDHs share a two-domain organization composed of a catalytic domain and a smaller flanking domain containing two CBS motifs	Pseudomonas aeruginosa

Synonyms

Synonyms	Comment	Organism
IMPDH	-	Pseudomonas aeruginosa
IMPDHpa	-	Pseudomonas aeruginosa
inosine-5'-monophosphate dehydrogenase	-	Pseudomonas aeruginosa

Temperature Optimum [°C]

Temperature Optimum [°C]	Temperature Optimum Maximum [°C]	Comment	Organism
30	-	assay at	Pseudomonas aeruginosa

Cofactor

Cofactor	Comment	Organism	Structure
NAD+	-	Pseudomonas aeruginosa

General Information

General Information	Comment	Organism
additional information	three conserved loops seem to be key players in the allosteric regulation of enzyme activity as they connect the tetramer-tetramer interface with the active site and show significant modification upon substrate binding, mechanism for the allosteric regulation of IMPDH through the CBS motifs. Conerved residue Asp199 is involved in the recognition of the ribose moiety of ATP in IMPDHpa. Structure analysis of wild-type and mutant enzymes, overview	Pseudomonas aeruginosa
physiological function	the key metabolic enzyme inosine-50-monophosphate dehydrogenase (IMPDH) occupies a central role in the de novo synthesis of guanosine nucleotides. The binding of Mg-ATP to Pseudomonas aeruginosa IMPDH and its crucial role in the regulation of the catalytic activity of the enzyme and its quaternary structure	Pseudomonas aeruginosa