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Literature summary for 1.1.1.307 extracted from

  • Fernandes, S.; Tuohy, M.G.; Murray, P.G.
    Xylose reductase from the thermophilic fungus Talaromyces emersonii: cloning and heterologous expression of the native gene (Texr) and a double mutant (TexrK271R + N273D) with altered coenzyme specificity (2009), J. Biosci., 34, 881-890.
    View publication on PubMed

Cloned(Commentary)

Cloned (Comment) Organism
expression in Escherichia coli Rasamsonia emersonii
expression of histidine-tagged wild type Texr and mutant Texr K271R/N273D in Escherichia coli BL21-Star DE3 Rasamsonia emersonii

Protein Variants

Protein Variants Comment Organism
K271R/N273D in the double mutant, affinity for NADPH decreases 3.1fold, while affinity for NADH remains relatively unchanged in comparison with the wild-type enzyme. The turnover number increases 1.6fold for the double mutant with NADH and decreases 3.2fold with NADPH relative to the wild-type enzyme. As a consequence, the catalytic efficiency of the double mutant (kcat/Km) increases 1.4fold with NADH and decreases 10.8fold with NADPH relative to the wild-type enzyme. Using the specificity constant (kcat/Km (NADH)/kcat/Km(NADPH)) the coenzyme preference for NADH is improved 16fold in the TeXR K271R/N273D double-mutant enzyme Rasamsonia emersonii
K271R/N273D catalytic efficiency of the double mutant increases 1.4fold with NADH and decreases 10.8fold with NADPH relative to the wild-type enzyme Rasamsonia emersonii

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
0.244
-
NADPH pH 6.5, 37°C, wild-type enzyme Rasamsonia emersonii
0.244
-
NADPH wild-type, pH 6.5, 37°C Rasamsonia emersonii
0.263
-
NADH pH 6.5, 37°C, wild-type enzyme Rasamsonia emersonii
0.263
-
NADH wild-type, pH 6.5, 37°C Rasamsonia emersonii
0.3
-
NADH pH 6.5, 37°C, mutant enzyme K271R/N273D Rasamsonia emersonii
0.3
-
NADH mutant K271R/N273, pH 6.5, 37°C Rasamsonia emersonii
0.747
-
NADPH pH 6.5, 37°C, mutant enzyme K271R/N273D Rasamsonia emersonii
0.747
-
NADPH mutant K271R/N273, pH 6.5, 37°C Rasamsonia emersonii
24.56
-
D-xylose pH 6.5, 37°C, cosubstrate: NADPH, wild-type enzyme Rasamsonia emersonii
24.6
-
D-xylose wild-type, cosubstrate NADPH, pH 6.5, 37°C Rasamsonia emersonii
76.06
-
D-xylose pH 6.5, 37°C, cosubstrate: NADH, mutant enzyme K271R/N273D Rasamsonia emersonii
76.1
-
D-xylose mutant K271R/N273, cosubstrate NADH, pH 6.5, 37°C Rasamsonia emersonii
76.5
-
D-xylose pH 6.5, 37°C, cosubstrate: NADPH, mutant enzyme K271R/N273D Rasamsonia emersonii
76.5
-
D-xylose mutant K271R/N273, cosubstrate NADPH, pH 6.5, 37°C Rasamsonia emersonii
89.4
-
D-xylose pH 6.5, 37°C, cosubstrate: NADH, wild-type enzyme Rasamsonia emersonii
89.4
-
D-xylose wild-type, cosubstrate NADH, pH 6.5, 37°C Rasamsonia emersonii

Molecular Weight [Da]

Molecular Weight [Da] Molecular Weight Maximum [Da] Comment Organism
36000
-
x * 36000, calculated, x * 40000, SDS-PAGE Rasamsonia emersonii
40000
-
x * 40000, wild-type and double-mutant (K271R/N273D) histidine-tagged protein, SDS-PAGE Rasamsonia emersonii
40000
-
x * 36000, calculated, x * 40000, SDS-PAGE Rasamsonia emersonii

Natural Substrates/ Products (Substrates)

Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
D-xylose + NADPH + H+ Rasamsonia emersonii expression of Texr is inducible by the same carbon sources responsible for the induction of genes encoding enzymes relevant to lignocellulose hydrolysis, suggesting a coordinated expression of intracellular and extracellular enzymes relevant to hydrolysis and metabolism of pentose sugars in Talaromyces emersonii in adaptation to its natural habitat. This indicates a potential advantage in survival and response to a nutrient-poor environment xylitol + NADP+
-
?

Organism

Organism UniProt Comment Textmining
Rasamsonia emersonii C5J3R6
-
-

Purification (Commentary)

Purification (Comment) Organism
-
Rasamsonia emersonii
recombinant protein Rasamsonia emersonii

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
D-xylose + NADH + H+
-
Rasamsonia emersonii xylitol + NAD+
-
?
D-xylose + NADH + H+ wild-type TeXR shows dual coenzyme specificity but is preferentially NADPH-dependent, with affinity for NADPH being 1.1fold higher than NADH and catalytic efficiency (kcat/Km) 24.5fold higher with NADPH as coenzyme. Affinity for xylose is 3.6fold higher with NADPH as coenzyme Rasamsonia emersonii xylitol + NAD+
-
?
D-xylose + NADPH + H+
-
Rasamsonia emersonii xylitol + NADP+
-
?
D-xylose + NADPH + H+ expression of Texr is inducible by the same carbon sources responsible for the induction of genes encoding enzymes relevant to lignocellulose hydrolysis, suggesting a coordinated expression of intracellular and extracellular enzymes relevant to hydrolysis and metabolism of pentose sugars in Talaromyces emersonii in adaptation to its natural habitat. This indicates a potential advantage in survival and response to a nutrient-poor environment Rasamsonia emersonii xylitol + NADP+
-
?
D-xylose + NADPH + H+ wild-type TeXR shows dual coenzyme specificity but is preferentially NADPH-dependent, with affinity for NADPH being 1.1fold higher than NADH and catalytic efficiency (kcat/Km) 24.5fold higher with NADPH as coenzyme. Affinity for xylose is 3.6fold higher with NADPH as coenzyme Rasamsonia emersonii xylitol + NADP+
-
?

Subunits

Subunits Comment Organism
? x * 40000, wild-type and double-mutant (K271R/N273D) histidine-tagged protein, SDS-PAGE Rasamsonia emersonii
? x * 36000, calculated, x * 40000, SDS-PAGE Rasamsonia emersonii

Synonyms

Synonyms Comment Organism
Texr
-
Rasamsonia emersonii

Temperature Optimum [°C]

Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
37
-
-
Rasamsonia emersonii
37
-
wild-type and double-mutant (K271R/N273D) protein Rasamsonia emersonii

Temperature Range [°C]

Temperature Minimum [°C] Temperature Maximum [°C] Comment Organism
30 60 active from 30°C to 60°C, wild-type and double-mutant (K271R/N273D) protein Rasamsonia emersonii

Temperature Stability [°C]

Temperature Stability Minimum [°C] Temperature Stability Maximum [°C] Comment Organism
60
-
retained 28% of activity at 60°C Rasamsonia emersonii

Turnover Number [1/s]

Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
15750
-
D-xylose pH 6.5, 37°C, cosubstrate: NADH, wild-type enzyme Rasamsonia emersonii
15750
-
NADH wild-type, pH 6.5, 37°C Rasamsonia emersonii
25110
-
D-xylose pH 6.5, 37°C, cosubstrate: NADH, mutant enzyme K271R/N273D Rasamsonia emersonii
25110
-
NADH mutant K271R/N273, pH 6.5, 37°C Rasamsonia emersonii
100900
-
D-xylose pH 6.5, 37°C, cosubstrate: NADPH, mutant enzyme K271R/N273D Rasamsonia emersonii
100900
-
NADPH mutant K271R/N273, pH 6.5, 37°C Rasamsonia emersonii
324000
-
D-xylose pH 6.5, 37°C, cosubstrate: NADPH, wild-type enzyme Rasamsonia emersonii
324000
-
NADPH wild-type, pH 6.5, 37°C Rasamsonia emersonii

pH Optimum

pH Optimum Minimum pH Optimum Maximum Comment Organism
6.5
-
-
Rasamsonia emersonii
6.5
-
wild-type and double-mutant (K271R/N273D) protein Rasamsonia emersonii

pH Range

pH Minimum pH Maximum Comment Organism
5 8
-
Rasamsonia emersonii
5 8 active from pH 5 to pH 8, wild-type and double-mutant (K271R/N273D) protein Rasamsonia emersonii

Cofactor

Cofactor Comment Organism Structure
NADH catalytic efficiency is 24.5fold higher with NADPH as coenzyme than with NADH Rasamsonia emersonii
NADPH wild-type TeXR shows dual coenzyme specificity but is preferentially NADPH-dependent, with affinity for NADPH being 1.1fold higher than NADH and catalytic efficiency (kcat/Km) 24.5fold higher with NADPH as coenzyme. Affinity for xylose is 3.6fold higher with NADPH as coenzyme. K271R/N273D double mutant displays an altered coenzyme preference with a 16fold improvement in NADH utilization relative to the wild type Rasamsonia emersonii
NADPH catalytic efficiency is 24.5fold higher with NADPH as coenzyme than with NADH. Affinity for xylose is 3.6fold higher with NADPH as coenzyme Rasamsonia emersonii

kcat/KM [mM/s]

kcat/KM Value [1/mMs-1] kcat/KM Value Maximum [1/mMs-1] Substrate Comment Organism Structure
59920
-
NADH pH 6.5, 37°C, wild-type enzyme Rasamsonia emersonii
59920
-
NADH wild-type, pH 6.5, 37°C Rasamsonia emersonii
83750
-
NADH pH 6.5, 37°C, mutant enzyme K271R/N273D Rasamsonia emersonii
83750
-
NADH mutant K271R/N273, pH 6.5, 37°C Rasamsonia emersonii
135500
-
NADPH pH 6.5, 37°C, mutant enzyme K271R/N273D Rasamsonia emersonii
135500
-
NADPH mutant K271R/N273, pH 6.5, 37°C Rasamsonia emersonii
1466000
-
NADPH pH 6.5, 37°C, wild-type enzyme Rasamsonia emersonii
1466000
-
NADPH wild-type, pH 6.5, 37°C Rasamsonia emersonii