Literature summary for 5.4.99.39 extracted from

Ito, R.; Masukawa, Y.; Nakada, C.; Amari, K.; Nakano, C.; Hoshino, T.
beta-Amyrin synthase from Euphorbia tirucalli. Steric bulk, not the Pi-electrons of Phe, at position 474 has a key role in affording the correct folding of the substrate to complete the normal polycyclization cascade (2014), Org. Biomol. Chem., 12, 3836-3846 .

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Cloned(Commentary)

Cloned (Comment)	Organism
recombinant expression of wild-type and mutant enzymes in Saccharomyces cerevisiae strain GIL77	Euphorbia tirucalli

Protein Variants

Protein Variants	Comment	Organism
F474A	site-directed mutagenesis, the mutant produces significantly larger amounts of the bicyclic products and a decreased amount of beta-amyrin compared to wild-type. The mutant variant produces (9betaH)-polypoda-7,13,17,21-tetraen-3beta-ol and (9betaH)-polypoda-8(26),13,17,21-tetraen-3beta-ol, which are generated from a chair-boat folding conformation	Euphorbia tirucalli
F474G	site-directed mutagenesis, the mutant produces significantly larger amounts of the bicyclic products and a decreased amount of beta-amyrin compared to wild-type	Euphorbia tirucalli
F474H	site-directed mutagenesis	Euphorbia tirucalli
F474L	site-directed mutagenesis, the mutant shows a significantly decreased production of bicyclic compounds, and in turn exhibits a higher production of beta-amyrin compared to wild-type	Euphorbia tirucalli
F474M	site-directed mutagenesis, the mutant shows a significantly decreased production of bicyclic compounds, and in turn exhibits a higher production of beta-amyrin compared to wild-type	Euphorbia tirucalli
F474T	site-directed mutagenesis	Euphorbia tirucalli
F474V	site-directed mutagenesis, the mutant shows a significantly decreased production of bicyclic compounds, and in turn exhibits a higher production of beta-amyrin compared to wild-type	Euphorbia tirucalli
F474W	site-directed mutagenesis	Euphorbia tirucalli
F474Y	site-directed mutagenesis	Euphorbia tirucalli

Natural Substrates/ Products (Substrates)

Natural Substrates	Organism	Comment (Nat. Sub.)	Natural Products	Comment (Nat. Pro.)	Rev.	Reac.
(3S)-2,3-epoxy-2,3-dihydrosqualene	Euphorbia tirucalli	-	beta-amyrin	-	r

Organism

Organism	UniProt	Comment	Textmining
Euphorbia tirucalli	Q401R6	-	-

Reaction

Reaction	Comment	Organism	Reaction ID
(3S)-2,3-epoxy-2,3-dihydrosqualene = beta-amyrin	cyclization pathway of (3S)-2,3-oxidosqualene to generate beta-amyrin triterpene via bicyclic, malabaricanyl, dammarenyl, baccharenyl, lupanyl, and oleanyl cation intermediates, detailed mechanism overview	Euphorbia tirucalli	a

Substrates and Products (Substrate)

Substrates	Comment Substrates	Organism	Products	Comment (Products)	Rev.	Reac.
(3S)-2,3-epoxy-2,3-dihydrosqualene	-	Euphorbia tirucalli	beta-amyrin	-	r
additional information	GC-MS and NMR spectroscopic product and metabolites identification from reactions of wild-type and mutant enzymes, overview	Euphorbia tirucalli	?	-	?

Synonyms

Synonyms	Comment	Organism
beta-amyrin cyclase	-	Euphorbia tirucalli
EtAS	-	Euphorbia tirucalli

General Information

General Information	Comment	Organism
evolution	residue F474 of Euphorbia tirucalli beta-amyrin cyclase is highly conserved in the superfamily of oxidosqualene cyclases	Euphorbia tirucalli
malfunction	enzyme mutant F474A and F474G produces significantly larger amounts of the bicyclic products and a decreased amount of beta-amyrin compared to wild-type. The mutant variant F474A produces (9betaH)-polypoda-7,13,17,21-tetraen-3beta-ol and (9betaH)-polypoda-8(26),13,17,21-tetraen-3beta-ol, which are generated from a chair-boat folding conformation. Substitutions with aliphatic amino acids lacking Pi-electrons such as Val, Leu, and Met lead to a significantly decreased production of bicyclic compounds, and in turn exhibit a higher production of beta-amyrin	Euphorbia tirucalli
additional information	catalytically important residue F474 residue is located near the B-ring formation site. The major role of Phe474 is not to stabilize the transient cation via cation-Pi interaction, but is to confer the appropriate steric bulk near the B-ring formation site, leading to the completion of the normal polycyclization pathway without accumulation of abortive cyclization products	Euphorbia tirucalli

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Release 2023.1 (January 2023)