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Literature summary for 1.13.11.60 extracted from

  • Seo, M.J.; Shin, K.C.; An, J.U.; Kang, W.R.; Ko, Y.J.; Oh, D.K.
    Characterization of a recombinant 7,8-linoleate diol synthase from Glomerella cingulate (2016), Appl. Microbiol. Biotechnol., 100, 3087-3099 .
    View publication on PubMed

Activating Compound

Activating Compound Comment Organism Structure
DMSO the whole enzyme activity is the highest using DMSO among the organic solvents tested, and its optimal concentration is 2.5% v/v Colletotrichum gloeosporioides

Cloned(Commentary)

Cloned (Comment) Organism
expression in Escherichia coli Colletotrichum gloeosporioides
phylogenetic tree, recombinant expression of wild-type and mutant His-tagged enzymes in Escherichia coli strain ER2566 Colletotrichum gloeosporioides

Protein Variants

Protein Variants Comment Organism
A891G site-directed mutagenesis, the mutant converts linoleic acid to 7,8-DiHODE as major product and 8,11-DiHODE as minor product, which is the similar pattern as the wild-type enzyme Colletotrichum gloeosporioides
C1038S site-directed mutagenesis, the mutant produces a low amount of 7,8-DiHODE with the accumulation of 8-HPODE Colletotrichum gloeosporioides
H1036A site-directed mutagenesis, the mutant produces a low amount of 7,8-DiHODE with the accumulation of 8-HPODE Colletotrichum gloeosporioides
H1036A/C1038S site-directed mutagenesis, the double mutant converts linoleic acid to only 8-HPODE Colletotrichum gloeosporioides
additional information comparison of substrate specificities of wild-type an d mutant enzymes, overview Colletotrichum gloeosporioides
N895D site-directed mutagenesis, the mutant produces a reduced amount of 8-HPODE, compared to wild-type, without the accumulation of DiHODE Colletotrichum gloeosporioides
N895L site-directed mutagenesis, the mutant converts linoleic acid to 7,8-DiHODE as major product and 8,11-DiHODE as minor product, which is the similar pattern as the wild-type enzyme Colletotrichum gloeosporioides
N895Q site-directed mutagenesis, the mutant converts linoleic acid to 7,8-DiHODE as major product and 8,11-DiHODE as minor product, which is the similar pattern as the wild-type enzyme Colletotrichum gloeosporioides
Q898E site-directed mutagenesis, the mutant converts linoleic acid to 8,11-DiHODE as major product and 7,8-DiHODE as minor product Colletotrichum gloeosporioides
Q898L site-directed mutagenesis, the mutant converts linoleic acid to 8,11-DiHODE as major product and 7,8-DiHODE as minor product Colletotrichum gloeosporioides
Y380F site-directed mutagenesis, the mutant enzyme produces trace amount of 7,8-DiHODE without the accumulation of 8-HPODE when incubated with linoleic acid as substrate Colletotrichum gloeosporioides

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
0.0193
-
linoleate N-terminal 8-dioxygenase domain, pH 6.5, 40°C Colletotrichum gloeosporioides
0.0193
-
(8R,9Z,12Z)-8-hydroperoxyoctadeca-9,12-dienoate recombinant enzyme, whole enzyme activity, pH 6.5, 40°C Colletotrichum gloeosporioides
0.022
-
linoleate bifunctional enzyme, pH 6.5, 40°C Colletotrichum gloeosporioides
0.0264
-
alpha-linolenate N-terminal 8-dioxygenase domain, pH 6.5, 40°C Colletotrichum gloeosporioides
0.0264
-
alpha-linolenate recombinant enzyme, whole enzyme activity, pH 6.5, 40°C Colletotrichum gloeosporioides
0.0293
-
oleate N-terminal 8-dioxygenase domain, pH 6.5, 40°C Colletotrichum gloeosporioides
0.0293
-
oleate recombinant enzyme, whole enzyme activity, pH 6.5, 40°C Colletotrichum gloeosporioides
0.0305
-
palmitoleate N-terminal 8-dioxygenase domain, pH 6.5, 40°C Colletotrichum gloeosporioides
0.0305
-
palmitoleate recombinant enzyme, whole enzyme activity, pH 6.5, 40°C Colletotrichum gloeosporioides
0.0319
-
eicosadienoic acid N-terminal 8-dioxygenase domain, pH 6.5, 40°C Colletotrichum gloeosporioides
0.0319
-
eicosadienoate recombinant enzyme, N-terminal domain activity, pH 6.5, 40°C Colletotrichum gloeosporioides
0.0324
-
eicosenoic acid N-terminal 8-dioxygenase domain, pH 6.5, 40°C Colletotrichum gloeosporioides
0.0324
-
eicosenoate recombinant enzyme, N-terminal domain activity, pH 6.5, 40°C Colletotrichum gloeosporioides
0.0333
-
oleate bifunctional enzyme, pH 6.5, 40°C Colletotrichum gloeosporioides
0.0358
-
arachidonic acid N-terminal 8-dioxygenase domain, pH 6.5, 40°C Colletotrichum gloeosporioides
0.0358
-
arachidonate recombinant enzyme, N-terminal domain activity, pH 6.5, 40°C Colletotrichum gloeosporioides
0.0368
-
dihomo-gamma-linolenic acid N-terminal 8-dioxygenase domain, pH 6.5, 40°C Colletotrichum gloeosporioides
0.0369
-
dihomo-gamma-linolenate recombinant enzyme, N-terminal domain activity, pH 6.5, 40°C Colletotrichum gloeosporioides
0.0371
-
alpha-linolenate bifunctional enzyme, pH 6.5, 40°C Colletotrichum gloeosporioides
0.048
-
palmitoleate bifunctional enzyme, pH 6.5, 40°C Colletotrichum gloeosporioides

Molecular Weight [Da]

Molecular Weight [Da] Molecular Weight Maximum [Da] Comment Organism
510000
-
gel filtration Colletotrichum gloeosporioides
510000
-
recombinant enzyme, gel filtration Colletotrichum gloeosporioides

Natural Substrates/ Products (Substrates)

Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
linoleate + O2 Colletotrichum gloeosporioides
-
(8R,9Z,12Z)-8-hydroperoxyoctadeca-9,12-dienoate
-
?

Organic Solvent Stability

Organic Solvent Comment Organism
DMSO maximum activity in presence of 2.5% DMSO Colletotrichum gloeosporioides

Organism

Organism UniProt Comment Textmining
Colletotrichum gloeosporioides A0A0S2SWE4 i.e. Colletotrichum gloeosporioides
-
Colletotrichum gloeosporioides A0A0S2SWE4 bifunctional enzyme, the second domain isomerizes the product (8R,9Z,12Z)-8-hydroperoxy-9,12-octadecadienoate to (7S,8S,9Z,12Z)-5,8-dihydroxyoctadeca-9,12-dienoate, cf. EC 5.4.4.6
-
Colletotrichum gloeosporioides KACC 40961 A0A0S2SWE4 i.e. Colletotrichum gloeosporioides
-

Purification (Commentary)

Purification (Comment) Organism
recombinant His-tagged wild-type and mutant enzymes 8fold from Escherichia coli strain ER2566 by nickel affinity chromatography Colletotrichum gloeosporioides

Source Tissue

Source Tissue Comment Organism Textmining
mycelium
-
Colletotrichum gloeosporioides
-

Specific Activity [micromol/min/mg]

Specific Activity Minimum [µmol/min/mg] Specific Activity Maximum [µmol/min/mg] Comment Organism
0.39
-
purified recombinant enzyme, whole enzyme activity, substrate palmitoleate, pH 6.5, 40°C Colletotrichum gloeosporioides
0.48
-
purified recombinant enzyme, whole enzyme activity, substrate oleate, pH 6.5, 40°C Colletotrichum gloeosporioides
0.75
-
purified recombinant enzyme, whole enzyme activity, substrate alpha-linolenate, pH 6.5, 40°C Colletotrichum gloeosporioides
0.87
-
pH 6.5, 40°C Colletotrichum gloeosporioides
0.87
-
purified recombinant enzyme, whole enzyme activity, substrate (8R,9Z,12Z)-8-hydroperoxyoctadeca-9,12-dienoate, pH 6.5, 40°C Colletotrichum gloeosporioides
1.17
-
purified recombinant enzyme, N-terminal domain activity, substrate arachidate, pH 6.5, 40°C Colletotrichum gloeosporioides
1.18
-
purified recombinant enzyme, N-terminal domain activity, substrate dihomo-gamma-linolenate, pH 6.5, 40°C Colletotrichum gloeosporioides
1.2
-
purified recombinant enzyme, N-terminal domain activity, substrate eicosadienoate, pH 6.5, 40°C Colletotrichum gloeosporioides
1.39
-
purified recombinant enzyme, N-terminal domain activity, substrate eicosenoate, pH 6.5, 40°C Colletotrichum gloeosporioides

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
alpha-linolenate + O2
-
Colletotrichum gloeosporioides (8R,9Z,12Z)-8-hydroperoxy-9,12,15-octadecatrienoate
-
?
alpha-linolenate + O2 via (9Z,12Z,15Z)-8-hydroperoxyoctadeca-9,12,15-trienoic acid, best substrate with respect to whole enzyme activity Colletotrichum gloeosporioides (9Z,12Z,15Z)-7,8-dihydroxyoctadeca-9,12,15-trienoic acid
-
?
arachidonate + O2
-
Colletotrichum gloeosporioides (5Z,8Z,11Z,14Z)-8-hydroxyeicosa-5,8,11,14-tetraenoic acid
-
?
arachidonic acid + O2
-
Colletotrichum gloeosporioides 8-hydroxyarachidonic acid
-
?
dihomo-gamma-linolenate + O2
-
Colletotrichum gloeosporioides (8Z,11Z,14Z)-8-hydroxyeicosa-8,11,14-trienoic acid
-
?
dihomo-gamma-linolenic acid + O2
-
Colletotrichum gloeosporioides 8-hydroxydihomo-gamma-linolenic acid
-
?
eicosadienoate + O2
-
Colletotrichum gloeosporioides (11Z,14Z)-8-hydroxyeicosa-11,14-dienoic acid
-
?
eicosadienoic acid + O2
-
Colletotrichum gloeosporioides 8-hydroxyeicosadienoic acid
-
?
eicosenoate + O2
-
Colletotrichum gloeosporioides (11Z)-8-hydroxyeicosa-11-enoic acid
-
?
eicosenoic acid + O2
-
Colletotrichum gloeosporioides 8-hydroxyeicosenoic acid
-
?
linoleate + O2
-
Colletotrichum gloeosporioides (8R,9Z,12Z)-8-hydroperoxy-9,12-octadecadienoate
-
?
linoleate + O2
-
Colletotrichum gloeosporioides KACC 40961 (8R,9Z,12Z)-8-hydroperoxy-9,12-octadecadienoate
-
?
linoleate + O2
-
Colletotrichum gloeosporioides (8R,9Z,12Z)-8-hydroperoxyoctadeca-9,12-dienoate
-
?
linoleate + O2
-
Colletotrichum gloeosporioides (7S,8S)-7,8-dihydroxy-(9Z,12Z)-octadecadienoic acid
-
?
linoleate + O2
-
Colletotrichum gloeosporioides (8R,9Z,12Z)-8-hydroperoxyoctadeca-9,12-dienoic acid
-
?
additional information bifunctional enzyme, the second domain isomerizes the product (8R,9Z,12Z)-8-hydroperoxy-9,12-octadecadienoate to (7S,8S,9Z,12Z)-5,8-dihydroxyoctadeca-9,12-dienoate, cf. EC 5.4.4.6 Colletotrichum gloeosporioides ?
-
?
additional information the bifunctional enzyme converts linoleic acid to a product, identified as (9Z,12Z)-7,8-dihydroxyoctadeca-9,12-dienoic acid (7,8-DiHODE) by liquid chromatography-mass spectrometry/mass spectrometry (LC-MS/MS) and nuclear magnetic resonance (NMR) spectroscopy. The specific activity and catalytic efficiency (kcat/Km) of 7,8-diol synthase for the conversion of fatty acid to dihydroxy fatty acid follows the descending order linoleic acid, alpha-linolenic acid, oleic acid, and palmitoleic acid, indicating that the enzyme is a 7,8-linoleate diol synthase (7,8-LDS). Substrate specificity, overview. The reaction via (9Z,12Z)-8-hydroperoxyoctadeca-9,12-dienoic acid (8-HPODE) as an intermediate and the accumulation of 8-HPODE is due to a higher 8-dioxygenase activity in the N-terminal domain than hydroperoxide isomerase activity in the C-terminal domain, EC 5.4.4.6. 8-HPODE is subsequently isomerized to (9Z,12Z)-7S,8S-dihydroxyoctadeca-9,12-dienoic acid (7,8-DiHODE) or (5R,8R,9Z,12Z)-5,8-dihydroxyoctadeca-9,12-dienoic acid (5,8-DiHODE) by the C-terminal hydroperoxide isomerase domain of diol synthase. No activity with conjugated linoleic acid, docosapentaenoic acid, and docosahexaenoic acid, and no isomerase activity and no whole enzyme acitivity, but dioxygenase activity with eicosenoic acid, eicosadienoic acid, dihomo-gamma-linolenic acid, and arachidonic acid Colletotrichum gloeosporioides ?
-
?
additional information bifunctional enzyme, the second domain isomerizes the product (8R,9Z,12Z)-8-hydroperoxy-9,12-octadecadienoate to (7S,8S,9Z,12Z)-5,8-dihydroxyoctadeca-9,12-dienoate, cf. EC 5.4.4.6 Colletotrichum gloeosporioides KACC 40961 ?
-
?
additional information the bifunctional enzyme converts linoleic acid to a product, identified as (9Z,12Z)-7,8-dihydroxyoctadeca-9,12-dienoic acid (7,8-DiHODE) by liquid chromatography-mass spectrometry/mass spectrometry (LC-MS/MS) and nuclear magnetic resonance (NMR) spectroscopy. The specific activity and catalytic efficiency (kcat/Km) of 7,8-diol synthase for the conversion of fatty acid to dihydroxy fatty acid follows the descending order linoleic acid, alpha-linolenic acid, oleic acid, and palmitoleic acid, indicating that the enzyme is a 7,8-linoleate diol synthase (7,8-LDS). Substrate specificity, overview. The reaction via (9Z,12Z)-8-hydroperoxyoctadeca-9,12-dienoic acid (8-HPODE) as an intermediate and the accumulation of 8-HPODE is due to a higher 8-dioxygenase activity in the N-terminal domain than hydroperoxide isomerase activity in the C-terminal domain, EC 5.4.4.6. 8-HPODE is subsequently isomerized to (9Z,12Z)-7S,8S-dihydroxyoctadeca-9,12-dienoic acid (7,8-DiHODE) or (5R,8R,9Z,12Z)-5,8-dihydroxyoctadeca-9,12-dienoic acid (5,8-DiHODE) by the C-terminal hydroperoxide isomerase domain of diol synthase. No activity with conjugated linoleic acid, docosapentaenoic acid, and docosahexaenoic acid, and no isomerase activity and no whole enzyme acitivity, but dioxygenase activity with eicosenoic acid, eicosadienoic acid, dihomo-gamma-linolenic acid, and arachidonic acid Colletotrichum gloeosporioides KACC 40961 ?
-
?
oleate + O2
-
Colletotrichum gloeosporioides (9Z)-8-hydroperoxyoctadec-9-enoate
-
?
oleate + O2
-
Colletotrichum gloeosporioides KACC 40961 (9Z)-8-hydroperoxyoctadec-9-enoate
-
?
oleate + O2 via (9Z)-8-hydroperoxyoctadeca-9-enoic acid Colletotrichum gloeosporioides (9Z)-7,8-dihydroxyoctadeca-9-enoic acid
-
?
palmitoleate + O2
-
Colletotrichum gloeosporioides (9Z)-8-hydroperoxyhexadec-9-enoate
-
?
palmitoleate + O2
-
Colletotrichum gloeosporioides KACC 40961 (9Z)-8-hydroperoxyhexadec-9-enoate
-
?
palmitoleate + O2 via (9Z)-dihydroperoxyhexadeca-9-enoate Colletotrichum gloeosporioides (9Z)-5,8-dihydroxyhexadeca-9-enoic acid
-
?

Subunits

Subunits Comment Organism
homotetramer 4 * 127000, recombinant His-tagged enzyme, SDS-PAGE, 4 * 127413, sequence calculation Colletotrichum gloeosporioides
More the enzyme diol synthase is a fusion protein, consisting of an N-terminal dioxygenase domain and a C-terminal cytochrome P450/hydroperoxide isomerase domain Colletotrichum gloeosporioides
trimer 3 * 127000, SDS-PAGE, 3 * 127413, calculated Colletotrichum gloeosporioides

Synonyms

Synonyms Comment Organism
8,11-hydroperoxide isomerase
-
Colletotrichum gloeosporioides

Temperature Optimum [°C]

Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
20
-
maximum of 8-diogenase reaction Colletotrichum gloeosporioides
20
-
N-terminal domain activity Colletotrichum gloeosporioides
40
-
maximum of whole enzyme Colletotrichum gloeosporioides
40
-
whole enzyme activity Colletotrichum gloeosporioides

Temperature Stability [°C]

Temperature Stability Minimum [°C] Temperature Stability Maximum [°C] Comment Organism
20 50 purified recombinant enzyme, 2 h, the activities of whole 7,8-LDS and 8-dioxygenase in the N-terminal domain decrease as temperature increases. The N-terminal domain demonstrates a greater thermal stability than the C-terminal domain. The whole enzyme activity presentes 72% of the initial activity at 40°C after 2 h, while 8-dioxygenase activity in the N-terminal domain is 90% Colletotrichum gloeosporioides

Turnover Number [1/s]

Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
4.86
-
palmitoleate bifunctional enzyme, pH 6.5, 40°C Colletotrichum gloeosporioides
5.62
-
oleate bifunctional enzyme, pH 6.5, 40°C Colletotrichum gloeosporioides
6.58
-
linoleate bifunctional enzyme, pH 6.5, 40°C Colletotrichum gloeosporioides
10.5
-
alpha-linolenate bifunctional enzyme, pH 6.5, 40°C Colletotrichum gloeosporioides
17.2
-
dihomo-gamma-linolenic acid N-terminal 8-dioxygenase domain, pH 6.5, 40°C Colletotrichum gloeosporioides
17.2
-
dihomo-gamma-linolenate recombinant enzyme, N-terminal domain activity, pH 6.5, 40°C Colletotrichum gloeosporioides
17.7
-
eicosadienoic acid N-terminal 8-dioxygenase domain, pH 6.5, 40°C Colletotrichum gloeosporioides
17.7
-
eicosadienoate recombinant enzyme, N-terminal domain activity, pH 6.5, 40°C Colletotrichum gloeosporioides
18.2
-
arachidonic acid N-terminal 8-dioxygenase domain, pH 6.5, 40°C Colletotrichum gloeosporioides
18.2
-
arachidonate recombinant enzyme, N-terminal domain activity, pH 6.5, 40°C Colletotrichum gloeosporioides
21.2
-
palmitoleate N-terminal 8-dioxygenase domain, pH 6.5, 40°C Colletotrichum gloeosporioides
21.2
-
palmitoleate recombinant enzyme, whole enzyme activity, pH 6.5, 40°C Colletotrichum gloeosporioides
22
-
eicosenoic acid N-terminal 8-dioxygenase domain, pH 6.5, 40°C Colletotrichum gloeosporioides
22
-
eicosenoate recombinant enzyme, N-terminal domain activity, pH 6.5, 40°C Colletotrichum gloeosporioides
24.7
-
oleate N-terminal 8-dioxygenase domain, pH 6.5, 40°C Colletotrichum gloeosporioides
24.7
-
oleate recombinant enzyme, whole enzyme activity, pH 6.5, 40°C Colletotrichum gloeosporioides
25.1
-
alpha-linolenate N-terminal 8-dioxygenase domain, pH 6.5, 40°C Colletotrichum gloeosporioides
25.2
-
alpha-linolenate recombinant enzyme, whole enzyme activity, pH 6.5, 40°C Colletotrichum gloeosporioides
35
-
linoleate N-terminal 8-dioxygenase domain, pH 6.5, 40°C Colletotrichum gloeosporioides
35
-
(8R,9Z,12Z)-8-hydroperoxyoctadeca-9,12-dienoate recombinant enzyme, whole enzyme activity, pH 6.5, 40°C Colletotrichum gloeosporioides

pH Optimum

pH Optimum Minimum pH Optimum Maximum Comment Organism
additional information
-
the maximum activity for the conversion of linoleic acid to (9Z,12Z)-8-hydroperoxyoctadeca-9,12-dienoic acid (8-HPODE) by 8-dioxygenase in the N-terminal domain of 7,8-LDS is observed at pH 7.0 and 20°C, whereas that for the conversion of linoleic acid to (9Z,12Z)-7,8-dihydroxyoctadeca-9,12-dienoic acid (7,8-DiHODE) by the whole enzyme is observed at pH 6.5 and 40°C Colletotrichum gloeosporioides
6.5
-
maximum of bifunctional enzyme Colletotrichum gloeosporioides
6.5
-
whole enzyme activity Colletotrichum gloeosporioides
7
-
maximum of 8-diogenase reaction Colletotrichum gloeosporioides
7
-
N-terminal domain activity Colletotrichum gloeosporioides

kcat/KM [mM/s]

kcat/KM Value [1/mMs-1] kcat/KM Value Maximum [1/mMs-1] Substrate Comment Organism Structure
100
-
palmitoleate bifunctional enzyme, pH 6.5, 40°C Colletotrichum gloeosporioides
170
-
oleate bifunctional enzyme, pH 6.5, 40°C Colletotrichum gloeosporioides
340
-
alpha-linolenate bifunctional enzyme, pH 6.5, 40°C Colletotrichum gloeosporioides
370
-
linoleate bifunctional enzyme, pH 6.5, 40°C Colletotrichum gloeosporioides
466.1
-
dihomo-gamma-linolenate recombinant enzyme, N-terminal domain activity, pH 6.5, 40°C Colletotrichum gloeosporioides
470
-
dihomo-gamma-linolenic acid N-terminal 8-dioxygenase domain, pH 6.5, 40°C Colletotrichum gloeosporioides
508.4
-
arachidonate recombinant enzyme, N-terminal domain activity, pH 6.5, 40°C Colletotrichum gloeosporioides
510
-
arachidonic acid N-terminal 8-dioxygenase domain, pH 6.5, 40°C Colletotrichum gloeosporioides
554.9
-
eicosadienoate recombinant enzyme, N-terminal domain activity, pH 6.5, 40°C Colletotrichum gloeosporioides
560
-
eicosadienoic acid N-terminal 8-dioxygenase domain, pH 6.5, 40°C Colletotrichum gloeosporioides
679
-
eicosenoate recombinant enzyme, N-terminal domain activity, pH 6.5, 40°C Colletotrichum gloeosporioides
680
-
eicosenoic acid N-terminal 8-dioxygenase domain, pH 6.5, 40°C Colletotrichum gloeosporioides
690
-
palmitoleate N-terminal 8-dioxygenase domain, pH 6.5, 40°C Colletotrichum gloeosporioides
695.1
-
palmitoleate recombinant enzyme, whole enzyme activity, pH 6.5, 40°C Colletotrichum gloeosporioides
840
-
oleate N-terminal 8-dioxygenase domain, pH 6.5, 40°C Colletotrichum gloeosporioides
843
-
oleate recombinant enzyme, whole enzyme activity, pH 6.5, 40°C Colletotrichum gloeosporioides
950
-
alpha-linolenate N-terminal 8-dioxygenase domain, pH 6.5, 40°C Colletotrichum gloeosporioides
954.54
-
alpha-linolenate recombinant enzyme, whole enzyme activity, pH 6.5, 40°C Colletotrichum gloeosporioides
1813.5
-
(8R,9Z,12Z)-8-hydroperoxyoctadeca-9,12-dienoate recombinant enzyme, whole enzyme activity, pH 6.5, 40°C Colletotrichum gloeosporioides
1820
-
linoleate N-terminal 8-dioxygenase domain, pH 6.5, 40°C Colletotrichum gloeosporioides