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Literature summary for 1.1.1.431 extracted from

  • Paidimuddala, B.; Mohapatra, S.B.; Gummadi, S.N.; Manoj, N.
    Crystal structure of yeast xylose reductase in complex with a novel NADP-DTT adduct provides insights into substrate recognition and catalysis (2018), FEBS J., 285, 4445-4464 .
    View publication on PubMed

Cloned(Commentary)

Cloned (Comment) Organism
recombinant expression of His-tagged enzyme in Escherichia coli strain Rosetta Debaryomyces nepalensis

Crystallization (Commentary)

Crystallization (Comment) Organism
purified xylose reductase (DnXR) in the apoform and as a ternary complex with a NADP-DTT adduct, covalent linkage between the C4N atom of the nicotinamide ring of the cosubstrate and the S1 sulfur atom of DTT and provides the first structural evidence for a protein mediated NADP-low-molecular-mass thiol adduct, the formation of the adduct is facilitated by an in-crystallo Michael addition of the DTT thiolate to the specific conformation of bound NADPH in the active site of DnXR. Hanging drop vapor diffusion method, mixing of 0.001 ml of 15 mg/ml protein in 20 mM Tris/HCl, pH 7.5, with 0.001 ml of reservoir solution containing 24% PEG 4000, 0.1 M sodium citrate, pH 6.2, and 0.15 M ammonium acetate, 20°C, for the complex crystals, 10 mM NADPH and 10 mM DTT are added to the protein solution and a reservoir solution containing 22% PEG 3350, 0.1 M HEPES, pH 7.5, and 0.2 M ammonium sulfate is used, X-ray diffraction structure determination and analysis at 1.7-2.0 A resolution, modeling Debaryomyces nepalensis

Inhibitors

Inhibitors Comment Organism Structure
DL-DTT inhibits DnXR via mixed inhibition mechanism, a covalent adduct is formed between the nicotinamide ring and enzyme-bound DTT (NADP-DTT). In the DnXR complex, DTT is bound in the substrate-binding pocket in an orientation expected for a four-carbon open chain sugar alcohol, reminiscent of a product-bound form. The tertiary structure has an eight stranded barrel (beta1-beta8) is flanked by eight helices (alpha1-alpha8). A beta-hairpin caps the N-terminal end of the barrel while the C-terminal end contains the conserved active site groove. The extended C-terminal region containing two helices reaches over the active site with the terminal loop that is stabilized by interactions with residues from loop 4. Sequence comparisons Debaryomyces nepalensis
meso-erythritol inhibits DnXR via mixed inhibition mechanism Debaryomyces nepalensis

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
additional information
-
additional information Michaelis-Menten and Lineweaver-Burk plots by linear and nonlinear regression fitting Debaryomyces nepalensis
5.5
-
D-erythrose recombinant His-tagged enzyme, pH 7.0, 45°C Debaryomyces nepalensis

Natural Substrates/ Products (Substrates)

Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
D-xylose + NADPH + H+ Debaryomyces nepalensis
-
xylitol + NADP+
-
r

Organism

Organism UniProt Comment Textmining
Debaryomyces nepalensis A0A0M4HL56
-
-

Purification (Commentary)

Purification (Comment) Organism
recombinant His-tagged enzyme from Escherichia coli strain Rosetta by nickel affinity chromatography Debaryomyces nepalensis

Reaction

Reaction Comment Organism Reaction ID
xylitol + NAD(P)+ = D-xylose + NAD(P)H + H+ structure-function analysis, overview Debaryomyces nepalensis

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
D-erythrose + NADPH + H+
-
Debaryomyces nepalensis erythritol + NADP+
-
r
D-xylose + NADPH + H+
-
Debaryomyces nepalensis xylitol + NADP+
-
r
L-threose + NADPH + H+
-
Debaryomyces nepalensis threitol + NADP+
-
r
additional information the enzyme displays the highest catalytic efficiency for L-threose, followed by D-erythrose. DnXR exhibits broad substrate specificity, with the highest catalytic efficiency for C5 sugars like arabinose, xylose, and ribose and a strict preference for cosubstrate NADPH Debaryomyces nepalensis ?
-
-

Subunits

Subunits Comment Organism
dimer
-
Debaryomyces nepalensis

Synonyms

Synonyms Comment Organism
DnXR
-
Debaryomyces nepalensis
xylose reductase
-
Debaryomyces nepalensis

Temperature Optimum [°C]

Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
45
-
-
Debaryomyces nepalensis

Turnover Number [1/s]

Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
165
-
D-erythrose recombinant His-tagged enzyme, pH 7.0, 45°C Debaryomyces nepalensis

pH Optimum

pH Optimum Minimum pH Optimum Maximum Comment Organism
7
-
-
Debaryomyces nepalensis

Cofactor

Cofactor Comment Organism Structure
NADP+
-
Debaryomyces nepalensis
NADPH enzyme DnXR shows strict dependence on NADPH, specific conformation of bound NADPH in the active site of DnXR, crystal structure analysis, structure-function analysis, overview. A large number of specific nonbonded interactions are involved in the binding. Among these, the side chain of Tyr212 stacks with the nicotinamide ring and hydrogen bond interactions between the side chains of Ser164, Asn165, and Gln186 with the carboxamide group orient the A-side of the nicotinamide group toward the substrate binding cavity Debaryomyces nepalensis

Ki Value [mM]

Ki Value [mM] Ki Value maximum [mM] Inhibitor Comment Organism Structure
1.8
-
DL-DTT recombinant His-tagged enzyme, pH 7.0, 45°C Debaryomyces nepalensis
1577
-
meso-erythritol recombinant His-tagged enzyme, pH 7.0, 45°C Debaryomyces nepalensis

General Information

General Information Comment Organism
evolution the xylose reductase (XR) belongs to the AKR2 family xylose reductase of aldo-keto reductase (AKR) superfamily Debaryomyces nepalensis
metabolism enzyme XR is the first enzyme in the xylose utilization pathway. Debaryomyces nepalensis, a nonpathogenic Saccharomycetes yeast can utilize both hexose and pentose sugars to produce polyols. DnXR is a key metabolic enzyme in the D-xylose utilization pathway Debaryomyces nepalensis
additional information the catalytic site architecture of AKRs includes a highly conserved tetrad of residues Asp42, Tyr47, Lys76, and His109 (DnXR numbering) lining the bottom of a deep open cavity Debaryomyces nepalensis
physiological function xylose reductase is a key enzyme in the conversion of xylose to xylitol, it catalyzes the conversion of carbonyl substrates into their respective alcohols Debaryomyces nepalensis

kcat/KM [mM/s]

kcat/KM Value [1/mMs-1] kcat/KM Value Maximum [1/mMs-1] Substrate Comment Organism Structure
30
-
D-erythrose recombinant His-tagged enzyme, pH 7.0, 45°C Debaryomyces nepalensis
138
-
L-threose recombinant His-tagged enzyme, pH 7.0, 45°C Debaryomyces nepalensis