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Literature summary 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)

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

Protein Variants

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

Inhibitors

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

KM Value [mM]

EC Number KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
1.1.1.9 0.027
-
 NAD+ native enzyme, at pH 7.0 and 30°C Rhizomucor pusillus
1.1.1.9 0.186
-
 NAD+ recombinant enzyme, at pH 7.0 and 30°C Rhizomucor pusillus
1.1.1.9 0.712
-
 NADP+ mutant enzyme D205A/I206R, at pH 7.0 and 30°C Rhizomucor pusillus
1.1.1.9 2.54
-
 NADP+ mutant enzyme D205A, at pH 7.0 and 30°C Rhizomucor pusillus
1.1.1.9 7.83
-
 xylitol native enzyme, at pH 7.0 and 30°C Rhizomucor pusillus
1.1.1.9 8.25
-
 D-xylulose at pH 7.0 and 30°C Rhizomucor pusillus
1.1.1.9 10.1
-
 xylitol recombinant enzyme, at pH 7.0 and 30°C Rhizomucor pusillus
1.1.1.9 10.2
-
 NADP+ recombinant enzyme, at pH 7.0 and 30°C Rhizomucor pusillus
1.1.1.9 13.5
-
 NAD+ mutant enzyme D205A, at pH 7.0 and 30°C Rhizomucor pusillus
1.1.1.9 19.8
-
 D-fructose at pH 7.0 and 30°C Rhizomucor pusillus
1.1.1.9 31.5
-
 D-sorbitol at pH 7.0 and 30°C Rhizomucor pusillus
1.1.1.9 34.3
-
 NAD+ mutant enzyme D205A/I206R, at pH 7.0 and 30°C Rhizomucor pusillus
1.1.1.9 66.7
-
 xylitol mutant enzyme D205A, with NAD+ as cosubstrate, at pH 7.0 and 30°C Rhizomucor pusillus
1.1.1.9 130
-
 xylitol mutant enzyme D205A/I206R, with NADP+ as cosubstrate, at pH 7.0 and 30°C Rhizomucor pusillus
1.1.1.9 149
-
 ribitol at pH 7.0 and 30°C Rhizomucor pusillus

Molecular Weight [Da]

EC Number Molecular Weight [Da] Molecular Weight Maximum [Da] Comment Organism
1.1.1.9 41000
-
 2 * 41000, SDS-PAGE Rhizomucor pusillus
1.1.1.9 87000
-
 gel filtration Rhizomucor pusillus

Natural Substrates/ Products (Substrates)

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

Organism

EC Number Organism UniProt Comment Textmining
1.1.1.9 Rhizomucor pusillus S6BFC0
-
-
1.1.1.9 Rhizomucor pusillus NBRC 4578 S6BFC0
-
-

Purification (Commentary)

EC Number Purification (Comment) Organism
1.1.1.9 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)

EC Number Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
1.1.1.9 D-fructose + NADH + H+
-
 Rhizomucor pusillus D-sorbitol + NAD+
-
 r
1.1.1.9 D-mannitol + NAD+ 2.6% activity compared to xylitol Rhizomucor pusillus ?
-
 ?
1.1.1.9 D-mannitol + NAD+ 2.6% activity compared to xylitol Rhizomucor pusillus NBRC 4578 ?
-
 ?
1.1.1.9 D-sorbitol + NAD+ 71.8% activity compared to xylitol Rhizomucor pusillus D-fructose + NADH + H+
-
 r
1.1.1.9 D-sorbitol + NAD+ 71.8% activity compared to xylitol Rhizomucor pusillus NBRC 4578 D-fructose + NADH + H+
-
 r
1.1.1.9 D-xylulose + NADH + H+
-
 Rhizomucor pusillus xylitol + NAD+
-
 r
1.1.1.9 D-xylulose + NADH + H+
-
 Rhizomucor pusillus NBRC 4578 xylitol + NAD+
-
 r
1.1.1.9 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 ?
-
 ?
1.1.1.9 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 ?
-
 ?
1.1.1.9 ribitol + NAD+ 60.1% activity compared to xylitol Rhizomucor pusillus ?
-
 ?
1.1.1.9 xylitol + NAD+ 100% activity Rhizomucor pusillus D-xylulose + NADH + H+
-
 r
1.1.1.9 xylitol + NAD+ 100% activity Rhizomucor pusillus NBRC 4578 D-xylulose + NADH + H+
-
 r
1.1.1.9 xylitol + NADP+ mutant enzyme D205A/I260R shows activity with NADP+ Rhizomucor pusillus D-xylulose + NADPH + H+
-
 r

Subunits

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

Synonyms

EC Number Synonyms Comment Organism
1.1.1.9 XDH
-
 Rhizomucor pusillus
1.1.1.9 XYL2
-
 Rhizomucor pusillus

Temperature Optimum [°C]

EC Number Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
1.1.1.9 60
-
-
 Rhizomucor pusillus

Temperature Stability [°C]

EC Number Temperature Stability Minimum [°C] Temperature Stability Maximum [°C] Comment Organism
1.1.1.9 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]

EC Number Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
1.1.1.9 0.185
-
 NADP+ recombinant enzyme, at pH 7.0 and 30°C Rhizomucor pusillus
1.1.1.9 0.33
-
 NADP+ mutant enzyme D205A, at pH 7.0 and 30°C Rhizomucor pusillus
1.1.1.9 0.4
-
 NADP+ mutant enzyme D205A/I206R, at pH 7.0 and 30°C Rhizomucor pusillus
1.1.1.9 1.16
-
 NAD+ mutant enzyme D205A/I206R, at pH 7.0 and 30°C Rhizomucor pusillus
1.1.1.9 5.18
-
 NAD+ mutant enzyme D205A, at pH 7.0 and 30°C Rhizomucor pusillus
1.1.1.9 6.35
-
 NAD+ recombinant enzyme, at pH 7.0 and 30°C Rhizomucor pusillus

pH Optimum

EC Number pH Optimum Minimum pH Optimum Maximum Comment Organism
1.1.1.9 9
-
-
 Rhizomucor pusillus

pH Range

EC Number pH Minimum pH Maximum Comment Organism
1.1.1.9 4 11 the enzyme loses its activity at pH higher than 11.0 and lower than 4.0 Rhizomucor pusillus

Cofactor

EC Number Cofactor Comment Organism Structure
1.1.1.9 NAD+ dependent on Rhizomucor pusillus

Expression

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

kcat/KM [mM/s]

EC Number kcat/KM Value [1/mMs-1] kcat/KM Value Maximum [1/mMs-1] Substrate Comment Organism Structure
1.1.1.9 0.018
-
 NADP+ recombinant enzyme, at pH 7.0 and 30°C Rhizomucor pusillus
1.1.1.9 0.03
-
 NAD+ mutant enzyme D205A/I206R, at pH 7.0 and 30°C Rhizomucor pusillus
1.1.1.9 0.13
-
 NADP+ mutant enzyme D205A, at pH 7.0 and 30°C Rhizomucor pusillus
1.1.1.9 0.38
-
 NAD+ mutant enzyme D205A, at pH 7.0 and 30°C Rhizomucor pusillus
1.1.1.9 0.56
-
 NADP+ mutant enzyme D205A/I206R, at pH 7.0 and 30°C Rhizomucor pusillus
1.1.1.9 32.36
-
 NAD+ recombinant enzyme, at pH 7.0 and 30°C Rhizomucor pusillus