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Literature summary for 1.1.1.358 extracted from

  • Cheng, P.; Wang, J.; Wu, Y.; Jiang, X.; Pei, X.; Su, W.
    Recombinant expression and molecular insights into the catalytic mechanism of an NADPH-dependent conjugated polyketone reductase for the asymmetric synthesis of (R)-pantolactone (2019), Enzyme Microb. Technol., 126, 77-85 .
    View publication on PubMed

Application

Application Comment Organism
synthesis the recombinant CduCPR from Candida dubliniensis exhibits potential application in the asymmetric synthesis of (R)-pantolactone Candida dubliniensis

Cloned(Commentary)

Cloned (Comment) Organism
gene CPR-C2, functional overexpression of N-terminally His6-tagged wild-type enzyme in Escherichia coli strain BL21(DE3), recombinant expression of His6-tagged enzyme mutants in Escherichia coli Candida dubliniensis

Protein Variants

Protein Variants Comment Organism
H128A site-directed mutagenesis, the mutant shows highly reduced activity compared to wild-type enzyme Candida dubliniensis
T30A site-directed mutagenesis, the mutant shows reduced activity compared to wild-type enzyme Candida dubliniensis
Y66A site-directed mutagenesis, inactive mutant Candida dubliniensis

Inhibitors

Inhibitors Comment Organism Structure
Ag+ complete inhibition at 1 mM Candida dubliniensis
Ca2+ 5.4% inhibition at 1 mM Candida dubliniensis
Co2+ 24.7% inhibition at 1 mM Candida dubliniensis
Cu2+ 70.3% inhibition at 1 mM Candida dubliniensis
EDTA 14.3% inhibition at 1 mM Candida dubliniensis
Fe2+ 33.2% inhibition at 1 mM Candida dubliniensis
Mg2+ 22.4% inhibition at 1 mM Candida dubliniensis
Mn2+ 43.2% inhibition at 1 mM Candida dubliniensis
Ni2+ 11.5% inhibition at 1 mM Candida dubliniensis
Pb2+ complete inhibition at 1 mM Candida dubliniensis
Zn2+ 89.2% inhibition at 1 mM Candida dubliniensis

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
0.484
-
2-dehydropantolactone pH 7.0, 30°C, recombinant wild-type enzyme Candida dubliniensis
3.5
-
2-dehydropantolactone pH 7.0, 30°C, recombinant mutant T30A Candida dubliniensis
20.4
-
2-dehydropantolactone pH 7.0, 30°C, recombinant mutant H128A Candida dubliniensis

Natural Substrates/ Products (Substrates)

Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
2-dehydropantolactone + NADPH + H+ Candida dubliniensis
-
(R)-pantolactone + NADP+
-
?
2-dehydropantolactone + NADPH + H+ Candida dubliniensis ATCC MYA-646
-
(R)-pantolactone + NADP+
-
?
2-dehydropantolactone + NADPH + H+ Candida dubliniensis CD36
-
(R)-pantolactone + NADP+
-
?
2-dehydropantolactone + NADPH + H+ Candida dubliniensis NCPF 3949
-
(R)-pantolactone + NADP+
-
?
2-dehydropantolactone + NADPH + H+ Candida dubliniensis NRRL Y-17841
-
(R)-pantolactone + NADP+
-
?
2-dehydropantolactone + NADPH + H+ Candida dubliniensis CBS 7987
-
(R)-pantolactone + NADP+
-
?

Organism

Organism UniProt Comment Textmining
Candida dubliniensis B9WJQ5
-
-
Candida dubliniensis ATCC MYA-646 B9WJQ5
-
-
Candida dubliniensis CBS 7987 B9WJQ5
-
-
Candida dubliniensis CD36 B9WJQ5
-
-
Candida dubliniensis NCPF 3949 B9WJQ5
-
-
Candida dubliniensis NRRL Y-17841 B9WJQ5
-
-

Purification (Commentary)

Purification (Comment) Organism
recombinant His6-tagged enzyme from Escherichia coli strain BL21(DE3) by nickel affinity chromatography and dialysis Candida dubliniensis

Reaction

Reaction Comment Organism Reaction ID
(R)-pantolactone + NADP+ = 2-dehydropantolactone + NADPH + H+ catalytic mechanism of (R)-pantolactone synthesis, overview. The conserved catalytic triad is formed by residues Thr30, Tyr66 and His128. Tyr66 functions as a proton donor following hydrogen transfer from NADPH. Thr30 and His128 are critical residues to bind and orient 2-dehydropantolactone (KPL). The phenolic hydroxyl group of Tyr66 forms the hydrogen bond with the C5 carbonyl oxygen of 2-dehydropantolactone. The HR of NADPH could just attack the si-face of the carbonyl group, which results in the formation of (R)-pantolactone. The alpha-amino group and the hydroxyl group of Thr30 form hydrogen bonds with the C4 carbonyl oxygen of KPL, and the epsilon-amino group of Lys33 formes the hydrogen bond with the O2 of KPL. In addition, the hydrophobic interaction of His128, Val165 and Phe302 with two methyl groups of KPL might facilitate the substrate binding Candida dubliniensis

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
2-dehydropantolactone + NADPH + H+
-
Candida dubliniensis (R)-pantolactone + NADP+
-
?
2-dehydropantolactone + NADPH + H+
-
Candida dubliniensis ATCC MYA-646 (R)-pantolactone + NADP+
-
?
2-dehydropantolactone + NADPH + H+
-
Candida dubliniensis CD36 (R)-pantolactone + NADP+
-
?
2-dehydropantolactone + NADPH + H+
-
Candida dubliniensis NCPF 3949 (R)-pantolactone + NADP+
-
?
2-dehydropantolactone + NADPH + H+
-
Candida dubliniensis NRRL Y-17841 (R)-pantolactone + NADP+
-
?
2-dehydropantolactone + NADPH + H+
-
Candida dubliniensis CBS 7987 (R)-pantolactone + NADP+
-
?
additional information enzyme CduCPR shows high catalytic activity and stereoselectivity using 2-dehydropantolactone (KPL) as the substrate. In a continuous feeding reaction, 200 mM ketopantolactone is reduced to (R)-pantolactone with 98% conversion and 99% enantiomeric excess within 2.0 h by the purified recombinant enzyme Candida dubliniensis ?
-
-
additional information enzyme CduCPR shows high catalytic activity and stereoselectivity using 2-dehydropantolactone (KPL) as the substrate. In a continuous feeding reaction, 200 mM ketopantolactone is reduced to (R)-pantolactone with 98% conversion and 99% enantiomeric excess within 2.0 h by the purified recombinant enzyme Candida dubliniensis ATCC MYA-646 ?
-
-
additional information enzyme CduCPR shows high catalytic activity and stereoselectivity using 2-dehydropantolactone (KPL) as the substrate. In a continuous feeding reaction, 200 mM ketopantolactone is reduced to (R)-pantolactone with 98% conversion and 99% enantiomeric excess within 2.0 h by the purified recombinant enzyme Candida dubliniensis CD36 ?
-
-
additional information enzyme CduCPR shows high catalytic activity and stereoselectivity using 2-dehydropantolactone (KPL) as the substrate. In a continuous feeding reaction, 200 mM ketopantolactone is reduced to (R)-pantolactone with 98% conversion and 99% enantiomeric excess within 2.0 h by the purified recombinant enzyme Candida dubliniensis NCPF 3949 ?
-
-
additional information enzyme CduCPR shows high catalytic activity and stereoselectivity using 2-dehydropantolactone (KPL) as the substrate. In a continuous feeding reaction, 200 mM ketopantolactone is reduced to (R)-pantolactone with 98% conversion and 99% enantiomeric excess within 2.0 h by the purified recombinant enzyme Candida dubliniensis NRRL Y-17841 ?
-
-
additional information enzyme CduCPR shows high catalytic activity and stereoselectivity using 2-dehydropantolactone (KPL) as the substrate. In a continuous feeding reaction, 200 mM ketopantolactone is reduced to (R)-pantolactone with 98% conversion and 99% enantiomeric excess within 2.0 h by the purified recombinant enzyme Candida dubliniensis CBS 7987 ?
-
-

Synonyms

Synonyms Comment Organism
CduCPR
-
Candida dubliniensis
CPR
-
Candida dubliniensis
CPR-C2
-
Candida dubliniensis
NADPH-dependent conjugated polyketone reductase
-
Candida dubliniensis

Temperature Optimum [°C]

Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
40 45 recombinant enzyme Candida dubliniensis

Temperature Range [°C]

Temperature Minimum [°C] Temperature Maximum [°C] Comment Organism
25 55 over 40% of maximal activity within this range Candida dubliniensis

Temperature Stability [°C]

Temperature Stability Minimum [°C] Temperature Stability Maximum [°C] Comment Organism
30
-
purified recombinant His-tagged enzyme, pH 7.0, 7 h, about 90% activity remaining Candida dubliniensis
40
-
purified recombinant His-tagged enzyme, pH 7.0, 7 h, about 50% activity remaining Candida dubliniensis
50
-
purified recombinant His-tagged enzyme, pH 7.0, 7 h, about 10% activity remaining Candida dubliniensis

Turnover Number [1/s]

Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
6.01
-
2-dehydropantolactone pH 7.0, 30°C, recombinant mutant H128A Candida dubliniensis
15.5
-
2-dehydropantolactone pH 7.0, 30°C, recombinant mutant T30A Candida dubliniensis
59
-
2-dehydropantolactone pH 7.0, 30°C, recombinant wild-type enzyme Candida dubliniensis

pH Optimum

pH Optimum Minimum pH Optimum Maximum Comment Organism
6.5
-
recombinant enzyme Candida dubliniensis

pH Range

pH Minimum pH Maximum Comment Organism
5.5 8.5 over 40% of maximal activity within this range Candida dubliniensis

Cofactor

Cofactor Comment Organism Structure
NADPH
-
Candida dubliniensis

General Information

General Information Comment Organism
evolution enzyme CduCPR belongs to the aldo-keto reductase superfamily Candida dubliniensis
additional information homology modeled structure of CduCPR and molecular docking analysis, overview Candida dubliniensis

kcat/KM [mM/s]

kcat/KM Value [1/mMs-1] kcat/KM Value Maximum [1/mMs-1] Substrate Comment Organism Structure
0.295
-
2-dehydropantolactone pH 7.0, 30°C, recombinant mutant H128A Candida dubliniensis
4.43
-
2-dehydropantolactone pH 7.0, 30°C, recombinant mutant T30A Candida dubliniensis
121.9
-
2-dehydropantolactone pH 7.0, 30°C, recombinant wild-type enzyme Candida dubliniensis