Any feedback?
Please rate this page
(literature.php)
(0/150)

BRENDA support

Literature summary extracted from

  • Lopez-Gallego, F.; Agger, S.A.; Abate-Pella, D.; Distefano, M.D.; Schmidt-Dannert, C.
    Sesquiterpene synthases Cop4 and Cop6 from Coprinus cinereus: catalytic promiscuity and cyclization of farnesyl pyrophosphate geometric isomers (2010), ChemBioChem, 11, 1093-1106.
    View publication on PubMedView publication on EuropePMC

Activating Compound

EC Number Activating Compound Comment Organism Structure
4.2.3.128 additional information increasing the reaction temperature to 37°C decreases the fidelity of Cop4. At this temperature Cop4 generates a relative larger fraction of products beta-cubebene, sativene, delta-cadinene and beta-copaene, that are derived from a cadinyl cation intermediate Coprinopsis cinerea
4.2.3.129 additional information increasing the reaction temperature to 37°C decreases the fidelity of Cop4. At this temperature Cop4 generates a relative larger fraction of products beta-cubebene, sativene, delta-cadinene and beta-copaene, that are derived from a cadinyl cation intermediate Coprinopsis cinerea

Cloned(Commentary)

EC Number Cloned (Comment) Organism
4.2.3.91 expressed in Escherichia coli BL21 (DE3) cells Coprinopsis cinerea
4.2.3.95 expressed in Escherichia coli BL21(DE3) cells Coprinopsis cinerea

Protein Variants

EC Number Protein Variants Comment Organism
4.2.3.95 N224D the mutant makes a small amount of (6S)-beta-bisabolene (18%) from (2Z,6E)-farnesyl diphosphate, the mutation does not affect the product profile of Cop6 with its normal substrate (2E,6E)-farnesyl diphosphate Coprinopsis cinerea

KM Value [mM]

EC Number KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
4.2.3.75 0.011
-
 (2E,6E)-farnesyl diphosphate pH 8.0, 30°C Coprinopsis cinerea
4.2.3.91 0.011
-
 (2E,6E)-farnesyl diphosphate pH 8.0, 30°C Coprinopsis cinerea
4.2.3.91 0.011
-
 (2E,6E)-farnesyl diphosphate pH not specified in the publication, at 37°C Coprinopsis cinerea
4.2.3.95 0.0076
-
 (2E,6E)-farnesyl diphosphate at 25°C, pH not specified in the publication Coprinopsis cinerea
4.2.3.95 0.0076
-
 (2E,6E)-farnesyl diphosphate at 25°C, pH 8.0 Coprinopsis cinerea
4.2.3.128 additional information
-
 additional information kinetic analysis, overview Coprinopsis cinerea
4.2.3.129 additional information
-
 additional information kinetic analysis, overview Coprinopsis cinerea

Metals/Ions

EC Number Metals/Ions Comment Organism Structure
4.2.3.75 K+ replacement of Mg2+ by Mn2+ results in loss of product beta-copaene. Presence of K+ decreases the (?)-germacrene D yield and instead increases the overall yield of tricyclic sesquiterpene olefins Coprinopsis cinerea
4.2.3.75 Mg2+
-
 Coprinopsis cinerea
4.2.3.75 Mn2+ replacement of Mg2+ by Mn2+ results in loss of product beta-copaene. In the presence of the Mn2+, Cop4 favors a reaction path that ends after one cyclization in (?)-germacrene D Coprinopsis cinerea
4.2.3.91 K+ replacement of Mg2+ by Mn2+ results in loss of product beta-copaene. Presence of K+ decreases the (-)-germacrene D yield and instead increases the overall yield of tricyclic sesquiterpene olefins Coprinopsis cinerea
4.2.3.91 Mg2+
-
 Coprinopsis cinerea
4.2.3.91 Mn2+ replacement of Mg2+ by Mn2+ results in loss of product beta-copaene. In the presence of the Mn2+, Cop4 favors a reaction path that ends after one cyclization in (?)-germacrene D Coprinopsis cinerea
4.2.3.127 Mg2+ dependent on, two consensus sequences - an aspartate rich DDXXD/E and a NSE/DTE motif - located at the entrance of the active site coordinate a trinuclear Mg2+ cluster. In reactions where Mg2+ is replaced with either Mn2+ or K+ is the disappearance of beta-copaene Coprinopsis cinerea
4.2.3.127 additional information in presence of the divalent cation Mn2+, Cop4 favors a reaction path that ends after one cyclization in (-)-germacrene D Coprinopsis cinerea
4.2.3.128 Mg2+ required, substitution of Mg2+ with Mn2+ as the divalent metal ion shifts the product profile of Cop4 to germacrene D, disfavoring subsequent ring closures that produce the cadinyl cation and its tricyclic descendents. Two consensus sequences - an aspartate rich DDXXD/E and a NSE/DTE motif - located at the entrance of the active site coordinate a trinuclear Mg2+ cluster that ligands the diphosphate moiety of the isoprenoid substrate, positions the isoprenyl chain in the binding pocket and triggers closure of the active site along with diphosphate cleavage to generate an initial transoid, allylic carbocation Coprinopsis cinerea
4.2.3.128 Mn2+ substitution of Mg2+ with Mn2+ as the divalent metal ion shifts the product profile of Cop4 to germacrene D, disfavoring subsequent ring closures that produce the cadinyl cation and its tricyclic descendents Coprinopsis cinerea
4.2.3.128 NaCl does not affect the product specificity of Cop4 significantly at 1 M Coprinopsis cinerea
4.2.3.129 Mg2+ required, substitution of Mg2+ with Mn2+ as the divalent metal ion shifts the product profile of Cop4 to germacrene D, disfavoring subsequent ring closures that produce the cadinyl cation and its tricyclic descendents. Two consensus sequences - an aspartate rich DDXXD/E and a NSE/DTE motif - located at the entrance of the active site coordinate a trinuclear Mg2+ cluster that ligands the diphosphate moiety of the isoprenoid substrate, positions the isoprenyl chain in the binding pocket and triggers closure of the active site along with diphosphate cleavage to generate an initial transoid, allylic carbocation Coprinopsis cinerea
4.2.3.129 Mn2+ substitution of Mg2+ with Mn2+ as the divalent metal ion shifts the product profile of Cop4 to germacrene D, disfavoring subsequent ring closures that produce the cadinyl cation and its tricyclic descendents Coprinopsis cinerea
4.2.3.129 NaCl does not affect the product specificity of Cop4 significantly at 1 M Coprinopsis cinerea

Natural Substrates/ Products (Substrates)

EC Number Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
4.2.3.127 (2E,6E)-farnesyl diphosphate Coprinopsis cinerea
-
 beta-copaene + diphosphate
-
 ?
4.2.3.128 (2E,6E)-farnesyl diphosphate Coprinopsis cinerea
-
 beta-cubebene + diphosphate
-
 ?
4.2.3.128 additional information Coprinopsis cinerea Cop4 is a catalytically promiscuous enzyme that cyclizes (2E,6E)-farnesyl diphosphate into multiple products, including (-)-germacrene D and cubebol. Cop 4 produces several volatile sesquiterpene products, including delta-cadinene as the major product, when expressed in Escherichia coli ?
-
 ?
4.2.3.129 (2E,6E)-farnesyl diphosphate Coprinopsis cinerea
-
 (+)-sativene + diphosphate
-
 ?

Organism

EC Number Organism UniProt Comment Textmining
4.2.3.75 Coprinopsis cinerea
-
-
-
4.2.3.91 Coprinopsis cinerea
-
-
-
4.2.3.95 Coprinopsis cinerea
-
-
-
4.2.3.127 Coprinopsis cinerea A8NU13
-
-
4.2.3.128 Coprinopsis cinerea A8NU13
-
-
4.2.3.129 Coprinopsis cinerea A8NU13
-
-

Purification (Commentary)

EC Number Purification (Comment) Organism
4.2.3.91 Talon resin metal affinity column chromatography Coprinopsis cinerea
4.2.3.95 Talon resin column chromatography Coprinopsis cinerea

Reaction

EC Number Reaction Comment Organism Reaction ID
4.2.3.127 (2E,6E)-farnesyl diphosphate = beta-copaene + diphosphate cyclization reaction mechanism, overview Coprinopsis cinerea
a
4.2.3.128 (2E,6E)-farnesyl diphosphate = beta-cubebene + diphosphate reaction mechanism, structure-function relationship, overview. The reaction mechanism for (E,E)-FPP cyclization by Cop4 involves the 1,10-cyclization of a cisoid neryl cation to form a (Z,E)-germacradienyl cation, which undergoes a 1,3-hydride shift to form an allylic carbocation that is either deprotonated to yield (-)-germacrene D, the major product of Cop4, or 1,6 cyclized to the bicyclic cadinyl cation Coprinopsis cinerea
a

Substrates and Products (Substrate)

EC Number Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
4.2.3.75 (2E,6E)-farnesyl diphosphate conversion of (E,E)-FPP proceeds via an (E,E)-germacradienyl carbocation Coprinopsis cinerea (-)-germacrene D + diphosphate main products at pH 8.0 are 29.3% (-)-germacrene D, 28.2% cubebol, 10.4% delta-cadinene, at pH 10.0 is 91% (-)-germacrene D, at pH 5.0 84% (-)-germacrene D plus 12% delta-cadinene ?
4.2.3.75 additional information Coprinus cinereus Cop4 additionally accepts (2Z,6E)-farnesyl diphosphate to produce 27.7% alpha-acoradiene, 10% amorpha-4,11-diene, and 13.3% cadina-4,11-diene Coprinopsis cinerea ?
-
 ?
4.2.3.91 (2E,6E)-farnesyl diphosphate conversion of (6E,6E)-farnesyl diphosphate proceeds via an (E,E)-germacradienyl carbocation Coprinopsis cinerea cubebol + diphosphate main products at pH 8.0 are 29.3% (-)-germacrene D, 28.2% cubebol, 10.4% delta-cadinene, at pH 10.0 is 91% (-)-germacrene D, at pH 5.0 84% (-)-germacrene D plus 12% delta-cadinene ?
4.2.3.91 (2E,6E)-farnesyl diphosphate + H2O
-
 Coprinopsis cinerea cubebol + diphosphate
-
 ?
4.2.3.91 additional information Cop4 is a catalytically promiscuous enzyme that cyclizes (E,E)-farnesyl diphophate into multiple products, including (-)-germacrene D and cubebol (major products). Conversion of (E,E)-farnesyl diphophate by Cop4 proceeds via an (E,E)-germacradienyl carbocation. Cop4 cyclizes (2Z,6E)-farnesyl diphophate via a (6S)-beta-bisabolyl cation intermediate to (6S)-beta-bisabolene. Cop4 converts E-geranyl diphophate into cyclic and acyclic monoterpenes. Cop4 also cyclizes nerolidyl diphosphate. Under alkaline (pH 10) or acidic (pH 5.0) conditions, Cop4 becomes a very selective germacrene D synthase Coprinopsis cinerea ?
-
 ?
4.2.3.91 additional information Coprinus cinereus Cop4 additionally accepts (2Z,6E)-farnesyl diphosphate to produce 27.7% alpha-acoradiene, 10% amorpha-4,11-diene, and 13.3% cadina-4,11-diene Coprinopsis cinerea ?
-
 ?
4.2.3.95 (2E,6E)-farnesyl diphosphate conversion of (2E,6E)-farnesyl diphosphate proceeds via (3R)-nerolidyl diphosphate, a (6R)-beta-bisabolyl carbocation, and a (7R)-cuprenyl cation Coprinopsis cinerea (-)-alpha-cuprenene + diphosphate 98.2% yield ?
4.2.3.95 (2E,6E)-farnesyl diphosphate conversion of (2E,6E)-farnesyl diphosphate proceeds via a (6R)-beta-bisabolyl carbocation Coprinopsis cinerea (-)-alpha-cuprenene + diphosphate 98.2% yield ?
4.2.3.95 additional information cyclization of (2Z,6E)-farnesyl diphosphate by Cop6 proceeds through opposite enantiomers of a beta-bisabolyl cation intermediate and results in alpha-acoradiene + amorpha-4,11-diene Coprinopsis cinerea ?
-
 ?
4.2.3.127 (2E,6E)-farnesyl diphosphate
-
 Coprinopsis cinerea beta-copaene + diphosphate
-
 ?
4.2.3.127 (2E,6E)-farnesyl diphosphate conversion of (E,E)-farnesyl diphosphate proceeds via an (E,E)-germacradienyl carbocation in the case of Cop4. Cyclization of all-trans-farnesyl diphosphate compared to the cyclization of the cis-trans isomer of farnesyl diphosphate serving as a surrogate for the secondary cisoid neryl cation intermediate generated by sesquiterpene synthases capable of isomerizing the C2-C3 bond of all-trans-farnesyl diphosphate. (Z,E)-FPP is cyclized via a (6S)-beta-bisabolene carbocation by Cop4 Coprinopsis cinerea beta-copaene + diphosphate
-
 ?
4.2.3.127 (E)-geranyl diphosphate the enzyme accepts (E)-geranyl diphosphate as a substrate, but the catalytic efficiency with the shorter prenyl-diphosphate substrate is lower compared to their longer farnesyl diphosphate substrate Coprinopsis cinerea ?
-
 ?
4.2.3.127 additional information Cop4 is a catalytically promiscuous enzyme that cyclizes (E,E)-farnesyl diphosphate into multiple products, including (-)-germacrene D and cubebol. But changing the pH of the reaction drastically alters the fidelity of Cop4 and makes it a highly selective enzyme Coprinopsis cinerea ?
-
 ?
4.2.3.128 (2E,6E)-farnesyl diphosphate
-
 Coprinopsis cinerea beta-cubebene + diphosphate
-
 ?
4.2.3.128 (2E,6E)-farnesyl diphosphate conversion of (E,E)-farnesyl diphosphate proceeds via an (E,E)-germacradienyl carbocation in the case of Cop4 Coprinopsis cinerea beta-cubebene + diphosphate
-
 ?
4.2.3.128 (2Z,6E)-farnesyl diphosphate conversion of (E,E)-farnesyl diphosphate proceeds via a (6S)-beta-bisabolene carbocation in the case of Cop4 Coprinopsis cinerea beta-cubebene + diphosphate
-
 ?
4.2.3.128 (E)-geraniol diphosphate Cop4 converts 30% of (E)-geraniol diphosphate into cyclic monoterpene products. Limonene is synthesized as the major cyclic monoterpene product. Limonene can be derived from either a cisoid, exo- or cisoid, endo-conformation of the initial geranyl cation of the initial geranyl cation. Exo-conformation yields (Z)-beta-ocimene and linalool, while the endoconformation would give (E)-beta-ocimene Coprinopsis cinerea ?
-
 ?
4.2.3.128 additional information Cop4 is a catalytically promiscuous enzyme that cyclizes (2E,6E)-farnesyl diphosphate into multiple products, including (-)-germacrene D and cubebol. Cop 4 produces several volatile sesquiterpene products, including delta-cadinene as the major product, when expressed in Escherichia coli Coprinopsis cinerea ?
-
 ?
4.2.3.129 (2E,6E)-farnesyl diphosphate
-
 Coprinopsis cinerea (+)-sativene + diphosphate
-
 ?
4.2.3.129 (2E,6E)-farnesyl diphosphate conversion of (E,E)-farnesyl diphosphate proceeds via an (E,E)-germacradienyl carbocation in the case of Cop4 Coprinopsis cinerea (+)-sativene + diphosphate
-
 ?
4.2.3.129 (2Z,6E)-farnesyl diphosphate conversion of (E,E)-farnesyl diphosphate proceeds via a (6S)-beta-bisabolene carbocation in the case of Cop4 Coprinopsis cinerea (+)-sativene + diphosphate
-
 ?

Subunits

EC Number Subunits Comment Organism
4.2.3.127 More structural homology modeling, hydrogen bond interactions and metal ion coordination in the diphosphate bound closed conformation of the Cop models, overview Coprinopsis cinerea
4.2.3.128 More structural modeling, structure-function relationship, overview. Cop4 has a large binding pocket in the open conformation Coprinopsis cinerea
4.2.3.129 More structural modeling, structure-function relationship, overview. Cop4 has a large binding pocket in the open conformation Coprinopsis cinerea

Synonyms

EC Number Synonyms Comment Organism
4.2.3.75 Cop4
-
 Coprinopsis cinerea
4.2.3.91 Cop4
-
 Coprinopsis cinerea
4.2.3.95 (-)-alpha-cuprenene synthase
-
 Coprinopsis cinerea
4.2.3.95 Cop6
-
 Coprinopsis cinerea
4.2.3.127 Cop4
-
 Coprinopsis cinerea
4.2.3.127 sesquiterpene synthase
-
 Coprinopsis cinerea
4.2.3.128 Cop4
-
 Coprinopsis cinerea
4.2.3.128 sesquiterpene synthase
-
 Coprinopsis cinerea
4.2.3.129 Cop4
-
 Coprinopsis cinerea
4.2.3.129 sesquiterpene synthase
-
 Coprinopsis cinerea

Temperature Range [°C]

EC Number Temperature Minimum [°C] Temperature Maximum [°C] Comment Organism
4.2.3.75 4 25 lowering the reaction temperature from 25°C to 4°C increases the selectivity of Cop4 for (?)-germacrene D and decreases the fraction of structurally unidentified sesquiterpene olefins by half Coprinopsis cinerea
4.2.3.75 37
-
 increasing the reaction temperature to 37°C decreases the fidelity of Cop4. At 37°C Cop4 generates a relative larger fraction of products beta-cubebene, sativene, delta-cadinene and beta-copaene that are derived from a cadinyl cation intermediate Coprinopsis cinerea
4.2.3.91 4 25 lowering the reaction temperature from 25°C to 4°C increases the selectivity of Cop4 for (-)-germacrene D and decreases the fraction of structurally unidentified sesquiterpene olefins by half Coprinopsis cinerea
4.2.3.91 37
-
 increasing the reaction temperature to 37°C decreases the fidelity of Cop4. At 37°C Cop4 generates a relative larger fraction of products beta-cubebene, sativene, delta-cadinene and beta-copaene that are derived from a cadinyl cation intermediate Coprinopsis cinerea
4.2.3.127 additional information
-
 lowering the reaction temperature from 25°C to 4°C increases the selectivity of Cop4 for (-)-germacrene D. Increasing the reaction temperature to 37°C has the opposite effect and decreases the fidelity of Cop4. At this temperature Cop4 generates a relative larger fraction of products beta-cubebene, sativene delta-cadinene, and beta-copaene that are derived from a cadinyl cation intermediate Coprinopsis cinerea

Turnover Number [1/s]

EC Number Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
4.2.3.75 0.012
-
 (2E,6E)-farnesyl diphosphate pH 8.0, 30°C Coprinopsis cinerea
4.2.3.91 0.012
-
 (2E,6E)-farnesyl diphosphate pH 8.0, 30°C Coprinopsis cinerea
4.2.3.91 0.012
-
 (2E,6E)-farnesyl diphosphate pH not specified in the publication, at 37°C Coprinopsis cinerea
4.2.3.95 0.67
-
 (2E,6E)-farnesyl diphosphate at 25°C, pH not specified in the publication Coprinopsis cinerea
4.2.3.95 0.67
-
 (2E,6E)-farnesyl diphosphate at 25°C, pH 8.0 Coprinopsis cinerea

pH Optimum

EC Number pH Optimum Minimum pH Optimum Maximum Comment Organism
4.2.3.75 additional information
-
 under both alkaline and acidic conditions Cop4 becomes a very selective enzyme with only one major product (?)-germacrene D compared to three major compounds produced under neutral reaction conditions. At pH 10, the cyclization reaction ends with the hydride shift and deprotonation of the germacradienyl cation to yield (?)-germacrene D, 91% of total sesquiterpene products, while at pH 5.0 cyclization can proceed via a 1,6 ring closure to also yield a small amount of delta-cadinene, 12% of total sesquiterpene products, in addition to (?)-germacrene D Coprinopsis cinerea
4.2.3.75 5
-
 at pH 5.0 cyclization can proceed via a 1,6 ring closure to also yield a small amount of delta-cadinene, 12% of total sesquiterpene products, in addition to (?)-germacrene D Coprinopsis cinerea
4.2.3.75 10
-
 at pH 10, the cyclization reaction ends with the hydride shift and deprotonation of the germacradienyl cation to yield (?)-germacrene D, 91% of total sesquiterpene products Coprinopsis cinerea
4.2.3.91 additional information
-
 under both alkaline and acidic conditions Cop4 becomes a very selective enzyme with only one major product (-)-germacrene D compared to three major compounds produced under neutral reaction conditions. At pH 10, the cyclization reaction ends with the hydride shift and deprotonation of the germacradienyl cation to yield (-)-germacrene D, 91% of total sesquiterpene products, while at pH 5.0 cyclization can proceed via a 1,6 ring closure to also yield a small amount of delta-cadinene, 12% of total sesquiterpene products, in addition to (?)-germacrene D Coprinopsis cinerea
4.2.3.91 5
-
 at pH 5.0 cyclization can proceed via a 1,6 ring closure to also yield a small amount of delta-cadinene, 12% of total sesquiterpene products, in addition to (-)-germacrene D Coprinopsis cinerea
4.2.3.91 10
-
 at pH 10, the cyclization reaction ends with the hydride shift and deprotonation of the germacradienyl cation to yield (-)-germacrene D, 91% of total sesquiterpene products Coprinopsis cinerea

pH Range

EC Number pH Minimum pH Maximum Comment Organism
4.2.3.75 additional information
-
 the catalytic fidelity of Cop4 can be drastically altered by varying reactions conditions Coprinopsis cinerea
4.2.3.91 additional information
-
 the catalytic fidelity of Cop4 can be drastically altered by varying reactions conditions Coprinopsis cinerea
4.2.3.127 additional information
-
 changing the pH of the reaction drastically alters the fidelity of Cop4 and makes it a highly selective enzyme Coprinopsis cinerea
4.2.3.128 additional information
-
 changing the pH of the reaction drastically alters the fidelity of Cop4 and makes it a highly selective enzyme. Increasing the reaction temperature to 37°C decreases the fidelity of Cop4. At this temperature Cop4 generates a relative larger fraction of products beta-cubebene, sativene, delta-cadinene and beta-copaene, that are derived from a cadinyl cation intermediate. The histidine side chain in the Cop4 loop, in particular, has a strong impact on the net charge of the loop at different pH values Coprinopsis cinerea
4.2.3.129 additional information
-
 changing the pH of the reaction drastically alters the fidelity of Cop4 and makes it a highly selective enzyme. Increasing the reaction temperature to 37°C decreases the fidelity of Cop4. At this temperature Cop4 generates a relative larger fraction of products beta-cubebene, sativene, delta-cadinene and beta-copaene, that are derived from a cadinyl cation intermediate. The histidine side chain in the Cop4 loop, in particular, has a strong impact on the net charge of the loop at different pH values Coprinopsis cinerea

General Information

EC Number General Information Comment Organism
4.2.3.128 additional information structural modeling, structure-function relationship, overview. Changing the pH of the reaction drastically alters the fidelity of Cop4 and makes it a highly selective enzyme Coprinopsis cinerea
4.2.3.129 additional information structural modeling, structure-function relationship, overview. Changing the pH of the reaction drastically alters the fidelity of Cop4 and makes it a highly selective enzyme Coprinopsis cinerea

kcat/KM [mM/s]

EC Number kcat/KM Value [1/mMs-1] kcat/KM Value Maximum [1/mMs-1] Substrate Comment Organism Structure
4.2.3.75 1
-
 (2E,6E)-farnesyl diphosphate pH 8.0, 30°C Coprinopsis cinerea
4.2.3.95 88
-
 (2E,6E)-farnesyl diphosphate at 25°C, pH not specified in the publication Coprinopsis cinerea
4.2.3.95 88
-
 (2E,6E)-farnesyl diphosphate at 25°C, pH 8.0 Coprinopsis cinerea