Literature summary for 4.2.3.128 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.

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Activating Compound

Activating Compound	Comment	Organism	Structure
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

KM Value [mM]

KM Value [mM]	KM Value Maximum [mM]	Substrate	Comment	Organism	Structure
additional information	-	additional information	kinetic analysis, overview	Coprinopsis cinerea

Metals/Ions

Metals/Ions	Comment	Organism	Structure
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
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
NaCl	does not affect the product specificity of Cop4 significantly at 1 M	Coprinopsis cinerea

Natural Substrates/ Products (Substrates)

Natural Substrates	Organism	Comment (Nat. Sub.)	Natural Products	Comment (Nat. Pro.)	Rev.	Reac.
(2E,6E)-farnesyl diphosphate	Coprinopsis cinerea	-	beta-cubebene + diphosphate	-	?
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	?	-	?

Organism

Organism	UniProt	Comment	Textmining
Coprinopsis cinerea	A8NU13	-	-

Reaction

Reaction	Comment	Organism	Reaction ID
(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)

Substrates	Comment Substrates	Organism	Products	Comment (Products)	Rev.	Reac.
(2E,6E)-farnesyl diphosphate	-	Coprinopsis cinerea	beta-cubebene + diphosphate	-	?
(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	-	?
(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	-	?
(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	?	-	?
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	?	-	?

Subunits

Subunits	Comment	Organism
More	structural modeling, structure-function relationship, overview. Cop4 has a large binding pocket in the open conformation	Coprinopsis cinerea

Synonyms

Synonyms	Comment	Organism
Cop4	-	Coprinopsis cinerea
sesquiterpene synthase	-	Coprinopsis cinerea

pH Range

pH Minimum	pH Maximum	Comment	Organism
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

General Information	Comment	Organism
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

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Release 2023.1 (January 2023)