Literature summary for 4.1.99.12 extracted from

Echt, S.; Bauer, S.; Steinbacher, S.; Huber, R.; Bacher, A.; Fischer, M.
Potential anti-infective targets in pathogenic yeasts: structure and properties of 3,4-dihydroxy-2-butanone 4-phosphate synthase of Candida albicans (2004), J. Mol. Biol., 341, 1085-1096.

Search for more literature for 4.1.99.12

Application

Application	Comment	Organism
drug development	the enzyme is a potential anti-infective target in the pathogenic yeast	Candida albicans

Cloned(Commentary)

Cloned (Comment)	Organism
functional expression of the synthetic gene in Escherichia coli strains XL1-Blue and M15	Candida albicans

Crystallization (Commentary)

Crystallization (Comment)	Organism
purified recombinant apoenzyme in complex with the substrate ribulose 5-phosphate, sitting drop vapour diffusion method, 0.003 ml of 17-34 mg/ml protein in 50 mM Tris-HCl, pH 7.5, is mixed with 0.001 ml of reservoir solution containing 85 mM sodium citrate, pH 5.0, and 17% PEG 8000, with or without 5 mM EDTA, equilibration against 0.3 ml reservoir solution, 0.003 ml of the complex solution is mixed with 0.001 ml of 90 mM Mes/NaOH, pH 6.0, containing 18% PEG 8000, addition of 2 mM D-ribulose 5-phosphate, 20°C, X-ray diffraction structure determination and analysis at 1.6-1.7 A resolution, molecular replacement, modelling	Candida albicans

Crystallization (Comment)

Organism

purified recombinant apoenzyme in complex with the substrate ribulose 5-phosphate, sitting drop vapour diffusion method, 0.003 ml of 17-34 mg/ml protein in 50 mM Tris-HCl, pH 7.5, is mixed with 0.001 ml of reservoir solution containing 85 mM sodium citrate, pH 5.0, and 17% PEG 8000, with or without 5 mM EDTA, equilibration against 0.3 ml reservoir solution, 0.003 ml of the complex solution is mixed with 0.001 ml of 90 mM Mes/NaOH, pH 6.0, containing 18% PEG 8000, addition of 2 mM D-ribulose 5-phosphate, 20°C, X-ray diffraction structure determination and analysis at 1.6-1.7 A resolution, molecular replacement, modelling

Candida albicans

Protein Variants

Protein Variants	Comment	Organism
C59A	site-directed mutagenesis, the mutant shows 70% of wild-type enzyme activity	Candida albicans
D92A	site-directed mutagenesis, inactive mutant	Candida albicans
E166A	site-directed mutagenesis, inactive mutant	Candida albicans
Q181R/Q183R	site-directed mutagenesis, construction of a synthetic gene, derived from orf 6.2440, with nucleotide exchanges at positions 414, 426, 477, 480, and 581	Candida albicans
Y87A	site-directed mutagenesis, the mutant shows 2% of wild-type enzyme activity	Candida albicans

Metals/Ions

Metals/Ions	Comment	Organism
Ca2+	not involved in catalysis, stabilize the enzyme	Candida albicans
Mg2+	essential for activity	Candida albicans
Zn2+	not involved in catalysis, stabilize the enzyme	Candida albicans

Molecular Weight [Da]

Molecular Weight [Da]	Molecular Weight Maximum [Da]	Comment	Organism
22530	-	2 * 22530, mass spectrometry, 2 * 22658, full-length enzyme, sequence calculation	Candida albicans
22658	-	2 * 22530, mass spectrometry, 2 * 22658, full-length enzyme, sequence calculation	Candida albicans
41000	-	recombinant enzyme, analytical ultracentrifugation, hydrodynamic studies	Candida albicans

Natural Substrates/ Products (Substrates)

Natural Substrates	Organism	Comment (Nat. Sub.)	Natural Products	Comment (Nat. Pro.)	Rev.	Reac.
D-ribulose 5-phosphate	Candida albicans	the enzyme is involved in riboflavin biosynthesis	formate + L-3,4-dihydroxy-2-butanone-4-phosphate	-	?

Organism

Organism	UniProt	Comment	Textmining
Candida albicans	Q5A3V6	wild-type enzyme; ATCC 10231D, orf 6.2440	-

Purification (Commentary)

Purification (Comment)	Organism
recombinant enzyme, expressed from the synthetic gene in Escherichia coli, to homogeneity by hydrophobic interaction chromatography, ultrafiltration, and gel filtration	Candida albicans

Reaction

Reaction	Comment	Organism	Reaction ID
D-ribulose 5-phosphate = formate + L-3,4-dihydroxybutan-2-one 4-phosphate	reaction mechanism, the enzyme possesses an essential acidic active-site loop	Candida albicans	a

Specific Activity [micromol/min/mg]

Specific Activity Minimum [µmol/min/mg]	Specific Activity Maximum [µmol/min/mg]	Comment	Organism
0.332	-	purified recombinant enzyme	Candida albicans

Substrates and Products (Substrate)

Substrates	Comment Substrates	Organism	Products	Comment (Products)	Rev.	Reac.
D-ribulose 5-phosphate	-	Candida albicans	formate + L-3,4-dihydroxy-2-butanone-4-phosphate	-	?
D-ribulose 5-phosphate	the enzyme is involved in riboflavin biosynthesis	Candida albicans	formate + L-3,4-dihydroxy-2-butanone-4-phosphate	-	?

Subunits

Subunits	Comment	Organism
dimer	2 * 22530, mass spectrometry, 2 * 22658, full-length enzyme, sequence calculation	Candida albicans
More	the enzyme possesses an essential acidic active-site loop, active site structure, structure comparisons, overview	Candida albicans

Synonyms

Synonyms	Comment	Organism
3,4-dihydroxy-2-butanone 4-phosphate synthase	-	Candida albicans
DHBPS	-	Candida albicans

Temperature Optimum [°C]

Temperature Optimum [°C]	Temperature Optimum Maximum [°C]	Comment	Organism
37	-	assay at	Candida albicans

pH Optimum

pH Optimum Minimum	pH Optimum Maximum	Comment	Organism
7.5	-	assay at	Candida albicans