Literature summary for 1.17.1.4 extracted from

Dietzel, U.; Kuper, J.; Doebbler, J.A.; Schulte, A.; Truglio, J.J.; Leimkuehler, S.; Kisker, C.
Mechanism of substrate and inhibitor binding of Rhodobacter capsulatus xanthine dehydrogenase (2009), J. Biol. Chem., 284, 8768-8776.

Cloned(Commentary)

Cloned (Comment)	Organism
expression of wild-type and mutant enzymes	Rhodobacter capsulatus

Crystallization (Commentary)

Crystallization (Comment)	Organism
purified recombinant wild-type enzyme and subunit beta mutants E232A and E232Q with or without molybdenum cofactor, hanging drop vapor diffusion method, 15 mg/ml protein in 50 mM Tris-HCl, pH 7.5, 1 mM EDTA, 200 mM NaCl,and 2.5 mM dithiothreitol are mixed 1 mM NAD+ and inhibitor pterin-6-aldehyde and with the reservoir solution containing 6-8 mM BaCl2, 6-8% polyethylene glycol 8000, 100 mM Tris-HCl, pH 8.3, 5-25mM dithiothreitol, and 3-4% isopropanol, whereas the EB232Q variant is mixed in a 1:2 ratio with the same reservoir solution, X-ray diffraction structure determination and analysis at 2.6-3.4 A resolution, overview	Rhodobacter capsulatus

Crystallization (Comment)

Organism

purified recombinant wild-type enzyme and subunit beta mutants E232A and E232Q with or without molybdenum cofactor, hanging drop vapor diffusion method, 15 mg/ml protein in 50 mM Tris-HCl, pH 7.5, 1 mM EDTA, 200 mM NaCl,and 2.5 mM dithiothreitol are mixed 1 mM NAD+ and inhibitor pterin-6-aldehyde and with the reservoir solution containing 6-8 mM BaCl2, 6-8% polyethylene glycol 8000, 100 mM Tris-HCl, pH 8.3, 5-25mM dithiothreitol, and 3-4% isopropanol, whereas the EB232Q variant is mixed in a 1:2 ratio with the same reservoir solution, X-ray diffraction structure determination and analysis at 2.6-3.4 A resolution, overview

Rhodobacter capsulatus

Protein Variants

Protein Variants	Comment	Organism
EB232Q	catalytically inactive active site mutant, inactive desulfo enzyme form	Rhodobacter capsulatus

Inhibitors

Inhibitors	Comment	Organism
allopurinol	i.e. 1-H-pyrazolo [3,4-d] pyrimidine-4-one	Rhodobacter capsulatus
oxypurinol	-	Rhodobacter capsulatus
pterin-6-aldehyde	competitive inhibition pattern, mechanism of inhibitor binding at the active site, overview	Rhodobacter capsulatus

Localization

Localization	Comment	Organism	GeneOntology No.	Textmining
cytosol	-	Rhodobacter capsulatus	5829	-

Natural Substrates/ Products (Substrates)

Natural Substrates	Organism	Comment (Nat. Sub.)	Natural Products	Comment (Nat. Pro.)	Rev.	Reac.
hypoxanthine + NAD+ + H2O	Rhodobacter capsulatus	-	xanthine + NADH + H+	-	?
xanthine + NAD+ + H2O	Rhodobacter capsulatus	-	urate + NADH + H+	-	?

Organism

Organism	UniProt	Comment	Textmining
Rhodobacter capsulatus	O54050	-	-

Purification (Commentary)

Purification (Comment)	Organism
recombinant wild-type and mutant enzymes with or without molybdenum cofactor by nickel affinity and anion exchange chromatography, followed by gel filtration	Rhodobacter capsulatus

Reaction

Reaction	Comment	Organism	Reaction ID
xanthine + NAD+ + H2O = urate + NADH + H+	the catalytic sequence of Rhodobacter capsulatus XDH is initiated by abstraction of a proton from the Mo-OH group by the highly conserved active site subunit B residue Glu730, followed by nucleophilic attack of the resulting Mo-O on the carbon center of the substrate (C-2 in hypoxanthine and C-8 in xanthine) and concomitant hydride transfer to the Mo-S of the molybdenum center, reaction and substrate binding mechanisms, overview	Rhodobacter capsulatus	a

Substrates and Products (Substrate)

Substrates	Comment Substrates	Organism	Products	Comment (Products)	Rev.
hypoxanthine + NAD+ + H2O	-	Rhodobacter capsulatus	xanthine + NADH + H+	-	?
hypoxanthine + NAD+ + H2O	mechanism of substrate binding at the active site, importance of beta subunit residue Glu232 for substrate positioning, overview. The oxygen atom at the C-6 position of both substrates is oriented toward ArgB-310 in the active site	Rhodobacter capsulatus	xanthine + NADH + H+	-	?
xanthine + NAD+ + H2O	-	Rhodobacter capsulatus	urate + NADH + H+	-	?
xanthine + NAD+ + H2O	mechanism of substrate binding at the active site, importance of beta subunit residue Glu232 for substrate positioning, overview. The oxygen atom at the C-6 position of both substrates is oriented toward ArgB-310 in the active site	Rhodobacter capsulatus	urate + NADH + H+	-	?

Subunits

Subunits	Comment	Organism
tetramer	(alphabeta)2 heterotetramer	Rhodobacter capsulatus

Synonyms

Synonyms	Comment	Organism
XDH	-	Rhodobacter capsulatus

Temperature Optimum [°C]

Temperature Optimum [°C]	Temperature Optimum Maximum [°C]	Comment	Organism
25	-	assay at	Rhodobacter capsulatus

pH Optimum

pH Optimum Minimum	pH Optimum Maximum	Comment	Organism
7.5	-	assay at	Rhodobacter capsulatus

Cofactor

Cofactor	Comment	Organism	Structure
NAD+	-	Rhodobacter capsulatus

Ki Value [mM]

Ki Value [mM]	Ki Value maximum [mM]	Inhibitor	Comment	Organism	Structure
0.001	-	oxypurinol	-	Rhodobacter capsulatus
0.1036	-	pterin-6-aldehyde	pH 7.5, 25°C	Rhodobacter capsulatus