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Literature summary for 1.1.1.9 extracted from

  • Yamasaki-Yashiki, S.; Komeda, H.; Hoshino, K.; Asano, Y.
    Molecular analysis of NAD+-dependent xylitol dehydrogenase from the zygomycetous fungus Rhizomucor pusillus and reversal of the coenzyme preference (2014), Biosci. Biotechnol. Biochem., 78, 1943-1953.
    View publication on PubMed

Cloned(Commentary)

Cloned (Comment) Organism
expressed in Escherichia coli BL21(DE3) cells Rhizomucor pusillus

Protein Variants

Protein Variants Comment Organism
D205A/I206R the mutant shows cofactor preference for NADP+ Rhizomucor pusillus

Inhibitors

Inhibitors Comment Organism Structure
Ag+ 9.2% residual activity at 1 mM Rhizomucor pusillus
Ca2+ complete inhibition at 1 mM Rhizomucor pusillus
Cd2+ complete inhibition at 1 mM Rhizomucor pusillus
Co2+ 6.8% residual activity at 1 mM Rhizomucor pusillus
Cu2+ complete inhibition at 1 mM Rhizomucor pusillus
Fe3+ 18% residual activity at 1 mM Rhizomucor pusillus
Mg2+ complete inhibition at 1 mM Rhizomucor pusillus
MgSO4 complete inhibition at 1 mM Rhizomucor pusillus
Mn2+ 8.4% residual activity at 1 mM Rhizomucor pusillus
Na+ complete inhibition at 1 mM Rhizomucor pusillus
Ni2+ 11.3% residual activity at 1 mM Rhizomucor pusillus
Pb2+ complete inhibition at 1 mM Rhizomucor pusillus
Zn2+ 44% residual activity at 1 mM Rhizomucor pusillus
ZnSO4 51.2% residual activity at 1 mM Rhizomucor pusillus

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
0.027
-
NAD+ native enzyme, at pH 7.0 and 30°C Rhizomucor pusillus
0.186
-
NAD+ recombinant enzyme, at pH 7.0 and 30°C Rhizomucor pusillus
0.712
-
NADP+ mutant enzyme D205A/I206R, at pH 7.0 and 30°C Rhizomucor pusillus
2.54
-
NADP+ mutant enzyme D205A, at pH 7.0 and 30°C Rhizomucor pusillus
7.83
-
xylitol native enzyme, at pH 7.0 and 30°C Rhizomucor pusillus
8.25
-
D-xylulose at pH 7.0 and 30°C Rhizomucor pusillus
10.1
-
xylitol recombinant enzyme, at pH 7.0 and 30°C Rhizomucor pusillus
10.2
-
NADP+ recombinant enzyme, at pH 7.0 and 30°C Rhizomucor pusillus
13.5
-
NAD+ mutant enzyme D205A, at pH 7.0 and 30°C Rhizomucor pusillus
19.8
-
D-fructose at pH 7.0 and 30°C Rhizomucor pusillus
31.5
-
D-sorbitol at pH 7.0 and 30°C Rhizomucor pusillus
34.3
-
NAD+ mutant enzyme D205A/I206R, at pH 7.0 and 30°C Rhizomucor pusillus
66.7
-
xylitol mutant enzyme D205A, with NAD+ as cosubstrate, at pH 7.0 and 30°C Rhizomucor pusillus
130
-
xylitol mutant enzyme D205A/I206R, with NADP+ as cosubstrate, at pH 7.0 and 30°C Rhizomucor pusillus
149
-
ribitol at pH 7.0 and 30°C Rhizomucor pusillus

Molecular Weight [Da]

Molecular Weight [Da] Molecular Weight Maximum [Da] Comment Organism
41000
-
2 * 41000, SDS-PAGE Rhizomucor pusillus
87000
-
gel filtration Rhizomucor pusillus

Natural Substrates/ Products (Substrates)

Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
D-xylulose + NADH + H+ Rhizomucor pusillus
-
xylitol + NAD+
-
r
D-xylulose + NADH + H+ Rhizomucor pusillus NBRC 4578
-
xylitol + NAD+
-
r
xylitol + NAD+ Rhizomucor pusillus 100% activity D-xylulose + NADH + H+
-
r
xylitol + NAD+ Rhizomucor pusillus NBRC 4578 100% activity D-xylulose + NADH + H+
-
r

Organism

Organism UniProt Comment Textmining
Rhizomucor pusillus S6BFC0
-
-
Rhizomucor pusillus NBRC 4578 S6BFC0
-
-

Purification (Commentary)

Purification (Comment) Organism
ammonium sulfate fractionation, DEAE Toyopearl chromatography, phenyl Toyopearl chromatography, Mono Q chromatography, Superdex 200 gel filtration, and Reactive Red column chromatography Rhizomucor pusillus

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
D-fructose + NADH + H+
-
Rhizomucor pusillus D-sorbitol + NAD+
-
r
D-mannitol + NAD+ 2.6% activity compared to xylitol Rhizomucor pusillus ?
-
?
D-mannitol + NAD+ 2.6% activity compared to xylitol Rhizomucor pusillus NBRC 4578 ?
-
?
D-sorbitol + NAD+ 71.8% activity compared to xylitol Rhizomucor pusillus D-fructose + NADH + H+
-
r
D-sorbitol + NAD+ 71.8% activity compared to xylitol Rhizomucor pusillus NBRC 4578 D-fructose + NADH + H+
-
r
D-xylulose + NADH + H+
-
Rhizomucor pusillus xylitol + NAD+
-
r
D-xylulose + NADH + H+
-
Rhizomucor pusillus NBRC 4578 xylitol + NAD+
-
r
additional information the enzyme shows less than 1% activity with erythritol, galactitol, D-arabitol, L-arabitol, and glycerol, and 3.1% activity with NADP+ compared to NAD+ Rhizomucor pusillus ?
-
?
additional information the enzyme shows less than 1% activity with erythritol, galactitol, D-arabitol, L-arabitol, and glycerol, and 3.1% activity with NADP+ compared to NAD+ Rhizomucor pusillus NBRC 4578 ?
-
?
ribitol + NAD+ 60.1% activity compared to xylitol Rhizomucor pusillus ?
-
?
xylitol + NAD+ 100% activity Rhizomucor pusillus D-xylulose + NADH + H+
-
r
xylitol + NAD+ 100% activity Rhizomucor pusillus NBRC 4578 D-xylulose + NADH + H+
-
r
xylitol + NADP+ mutant enzyme D205A/I260R shows activity with NADP+ Rhizomucor pusillus D-xylulose + NADPH + H+
-
r

Subunits

Subunits Comment Organism
homodimer 2 * 41000, SDS-PAGE Rhizomucor pusillus
homodimer 2 * 39185, calculated from amino acid sequence Rhizomucor pusillus

Synonyms

Synonyms Comment Organism
XDH
-
Rhizomucor pusillus
XYL2
-
Rhizomucor pusillus

Temperature Optimum [°C]

Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
60
-
-
Rhizomucor pusillus

Temperature Stability [°C]

Temperature Stability Minimum [°C] Temperature Stability Maximum [°C] Comment Organism
45 50 approximately 92% and 70% activities remain following incubations at 45 and 50°C, respectively. The enzyme completely loses its activity above 60°C Rhizomucor pusillus

Turnover Number [1/s]

Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
0.185
-
NADP+ recombinant enzyme, at pH 7.0 and 30°C Rhizomucor pusillus
0.33
-
NADP+ mutant enzyme D205A, at pH 7.0 and 30°C Rhizomucor pusillus
0.4
-
NADP+ mutant enzyme D205A/I206R, at pH 7.0 and 30°C Rhizomucor pusillus
1.16
-
NAD+ mutant enzyme D205A/I206R, at pH 7.0 and 30°C Rhizomucor pusillus
5.18
-
NAD+ mutant enzyme D205A, at pH 7.0 and 30°C Rhizomucor pusillus
6.35
-
NAD+ recombinant enzyme, at pH 7.0 and 30°C Rhizomucor pusillus

pH Optimum

pH Optimum Minimum pH Optimum Maximum Comment Organism
9
-
-
Rhizomucor pusillus

pH Range

pH Minimum pH Maximum Comment Organism
4 11 the enzyme loses its activity at pH higher than 11.0 and lower than 4.0 Rhizomucor pusillus

Cofactor

Cofactor Comment Organism Structure
NAD+ dependent on Rhizomucor pusillus

Expression

Organism Comment Expression
Rhizomucor pusillus the enzyme expression levels are increased with accumulation of xylitol up

kcat/KM [mM/s]

kcat/KM Value [1/mMs-1] kcat/KM Value Maximum [1/mMs-1] Substrate Comment Organism Structure
0.018
-
NADP+ recombinant enzyme, at pH 7.0 and 30°C Rhizomucor pusillus
0.03
-
NAD+ mutant enzyme D205A/I206R, at pH 7.0 and 30°C Rhizomucor pusillus
0.13
-
NADP+ mutant enzyme D205A, at pH 7.0 and 30°C Rhizomucor pusillus
0.38
-
NAD+ mutant enzyme D205A, at pH 7.0 and 30°C Rhizomucor pusillus
0.56
-
NADP+ mutant enzyme D205A/I206R, at pH 7.0 and 30°C Rhizomucor pusillus
32.36
-
NAD+ recombinant enzyme, at pH 7.0 and 30°C Rhizomucor pusillus