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Enzyme Nomenclature
Information on EC 4.2.3.61 - 5-epiaristolochene synthase
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4.2.3.61 5-epiaristolochene synthaseIUBMB Comments
Initial cyclization gives (+)-germacrene A in an enzyme bound form which is not released to the medium.
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The enzyme appears in viruses and cellular organisms
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SYNONYM 
ORGANISM
UNIPROT
COMMENTARY 
LITERATURE
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REACTION
REACTION DIAGRAM
COMMENTARY
ORGANISM
UNIPROT
LITERATURE
(2E,6E)-farnesyl diphosphate = (+)-5-epiaristolochene + diphosphate
-
-
-
-
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PATHWAY SOURCE
PATHWAYS
MetaCyc
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SYSTEMATIC NAME
IUBMB Comments
(2E,6E)-farnesyl-diphosphate diphosphate-lyase [(+)-5-epiaristolochene-forming]
Initial cyclization gives (+)-germacrene A in an enzyme bound form which is not released to the medium.
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SUBSTRATE
PRODUCT
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate)
(Substrate)
LITERATURE
(Substrate)
(Substrate)
COMMENTARY
(Product)
(Product)
LITERATURE
(Product)
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
r=reversible
ir=irreversible
?=not specified
(2E,6E)-farnesyl diphosphate
5-epi-aristolochene + diphosphate
-
-
-
?
(2E,6Z)-6-fluorofarnesyl diphosphate
(-)-1-fluorogermacrene A
-
the fluoro substitution at the C6 position of farnesyl diphosphate has negligible effects on enzyme binding, substrate orientation, diphosphate ionization, and the initial 1,10 ring closure catalyzed by TEAS
sole product, 58% yield
-
?
(2E,6E)-farnesyl diphosphate
(+)-5-epiaristolochene + diphosphate
-
single major sesquiterpene product
-
?
(2E,6E)-farnesyl diphosphate
(+)-5-epiaristolochene + diphosphate
-
-
-
-
?
(2E,6E)-farnesyl diphosphate
(+)-5-epiaristolochene + diphosphate
-
-
-
?
(2E,6E)-farnesyl diphosphate
(+)-5-epiaristolochene + diphosphate
-
-
(+)-5-epiaristolochene constitutes about 85% of hyxdrocarbon products
-
?
(2E,6E)-farnesyl diphosphate
(+)-5-epiaristolochene + diphosphate
-
-
about 79% of hydrocarbon product, plus about 6% of (-)-4-epi-eremophilene, 3.6% of (+)-germacrene A and 22 hydrocarbons contributing about 12% of TEAS sesquiterpene products. The pathway to the by-products starts with a 1,6 cyclization of the (Z,E)-farnesyl cation, followed by a 1,2 hydride shift leading to a syn-6,10 ring closure which generates an acoradilyl cation. Elimination of a proton from either C12 or C13 of the isopropylidene tail of the acoradilyl cation readily explains the formation of ?-acoradiene. Alternatively, a 2,11 ring closure followed by elimination of a proton at C2 is the likely pathway to (-)-alpha-cedrene. A 3,11 ring closure of the acoradilyl cation is the most common event, based on the relative abundance of isomerization products observed, and is followed by Wagner-Meerwein rearrangement and proton elimination from C15 to produce isoprezizaene, the dominant (Z,E)-farnesyl cation-derived product
-
?
(2E,6E)-farnesyl diphosphate
(+)-5-epiaristolochene + diphosphate
reaction proceeds via geracrene A as an intermediate. Proton donation by residue Y520 is responsible for the activation of germacrene A to a eudesmane cation
-
-
?
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METALS and IONS
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
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KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
0.0016
(2E,6E)-farnesyl diphosphate
-
fusion construct epiaristolochene synthase/farnesyl diphosphate synthase, pH 8.0, 30°C
0.0026
(2E,6E)-farnesyl diphosphate
-
fusion construct farnesyl diphosphate synthase/epiaristolochene synthase, pH 8.0, 30°C
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TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
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kcat/KM VALUE [1/mMs-1]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
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TEMPERATURE RANGE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
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ORGANISM
COMMENTARY
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
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SOURCE TISSUE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
SOURCE
additional information
additional information
no constitutive expression of EAS and capsidiol accumulation in shoots of rosette plants, but accumulation of transcripts can be induced in shoots by feeding of the tobacco hornworm, Manduca sexta
brenda
additional information
no constitutive expression of EAS and capsidiol accumulation in shoots of rosette plants, but accumulation of transcripts can be induced in shoots by feeding of the tobacco hornworm, Manduca sexta
brenda
additional information
no constitutive expression of EAS and capsidiol accumulation in shoots of rosette plants, but accumulation of transcripts can be induced in shoots by feeding of the tobacco hornworm, Manduca sexta
brenda
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UNIPROT
ENTRY NAME
ORGANISM
NO. OF AA
NO. OF TRANSM. HELICES
MOLECULAR WEIGHT[Da]
SOURCE
SEQUENCE
LOCALIZATION PREDICTION
The reliability class of the localization prediction ranges from 1 (very reliable) to 5 (not reliable).
The reliability class of the localization prediction ranges from 1 (very reliable) to 5 (not reliable). 5EAS1_NICAT
548
0
63167
Swiss-Prot
other Location (Reliability: 2)
5EAS2_NICAT
548
0
63190
Swiss-Prot
other Location (Reliability: 2)
5EAS3_NICAT
548
0
63155
Swiss-Prot
other Location (Reliability: 2)
5EAS4_NICAT
548
0
62897
Swiss-Prot
other Location (Reliability: 2)
5EAS_CAPAN
559
0
65098
Swiss-Prot
other Location (Reliability: 4)
5EAS_TOBAC
548
0
62987
Swiss-Prot
other Location (Reliability: 2)
A0A2G9GK65_9LAMI
555
0
64798
TrEMBL
other Location (Reliability: 3)
A0A2G9G9I1_9LAMI
125
0
14933
TrEMBL
other Location (Reliability: 3)
A0A2G9GK55_9LAMI
137
0
15743
TrEMBL
other Location (Reliability: 3)
A0A2G9HP72_9LAMI
180
0
20656
TrEMBL
other Location (Reliability: 2)
A0A2G9FWQ7_9LAMI
247
0
28533
TrEMBL
other Location (Reliability: 3)
A0A2G9GH43_9LAMI
129
0
15113
TrEMBL
other Location (Reliability: 2)
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PDB
SCOP
CATH
UNIPROT
ORGANISM
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CRYSTALLIZATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
to 2.2-2.8 A resolution, of TEAS alone and in complexes with two different substrate analogs. TEAS consists entirely of alpha-helices and short connecting loops and turns, and is organized into two structural domains. Two Mg2+ ions are coordinated on opposite sides of the entrance to the active site pocket, and constitute a diphosphate binding site. Asp301 coordinates a Mg2+ in the native TEAS structure, and the side chain carboxyl of Glu379 provides a longer range interaction. Asp305 provides an additional coordination bond in the enzyme with substrate analogs bound. Asp301 and Asp305 are part of a -DDXXD- sequence. Asp301 directly contacts Mg2+, whereas Asp302 demonstrates no direct metal coordination. The side chains of Asp444, Thr448, Glu452, and one water molecule the second coordinate Mg2+. In the native TEAS structure, the A-C and J-K loops and the residues NH2-terminal of residue 36 are disordered
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PROTEIN VARIANTS
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
Y520F
3% of wild-type catalytic efficiency, reaction prooduct is germacrene A
additional information
additional information
-
heterologous expression of EAS in transgenic rice does not interfere with the activities of endogenous squalene synthase or farnesyl diphosphatase. The induction of EAS enzyme activity is accompanied by an increase in EAS mRNA when challenged by the elicitor. The EAS ectopic expression in transgenic rice plants results in the synthesis of 5-epi-aristolochene in vivo upon elicitor treatment
additional information
-
construction of fusion proteins with farnesyl diphosphate synthase FPPS from Artemisia annua. The fusion enzymes produce epi-aristolochene from isopentenyl diphosphate through a coupled reaction. The Km values of FPPS and eAS for isopentenyl diphosphate and farnesyl diphosphate, respectively, are essentially the same for the single and fused enzymes. The bifunctional enzymes show a more efficient conversion of isopentenyl diphosphate to epi-aristolochene than the corresponding amount of single enzymes
additional information
engineering of a thermostable mutant by SCADS algorithm to suggest mutations based on side-chain interactions consistent with the nearby protein backbone, neighboring sidechains, and the local environment, and by replacement of solvent-exposed hydrophobic residues and the addition of salt bridges on the surface of the mutant enzyme. Contrary to wild-type, mutant is still active to 65°C but produces additional sesquiterpene products
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TEMPERATURE STABILITY
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
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PURIFICATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
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CLONED (Commentary)
ORGANISM
UNIPROT
LITERATURE
expression in Escherichia coli
expression in Petunia hybrida infiltrated with Agrobacterium tumefaciens harboring the enzyme gene
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EXPRESSION
ORGANISM
UNIPROT
LITERATURE
accumulation of transcripts can be induced in shoots by feeding of the tobacco hornworm, Manduca sexta
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APPLICATION
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
analysis
method for the recovery of full-length cDNAs from predicted terpene synthase genes containing introns. The approach utilizes Agrobacterium-mediated transient expression coupled with a reverse transcription-polydeoxyribonucleotide chain reaction assay to facilitate expression cloning of processed transcripts. Subsequent expression of intronless cDNAs in a suitable prokaryotic host provides for direct functional testing of the encoded gene product
synthesis
synthesis
-
construction of fusion proteins with farnesyl diphosphate synthase FPPS from Artemisia annua. The fusion enzymes produce epi-aristolochene from isopentenyl diphosphate through a coupled reaction. The Km values of FPPS and eAS for isopentenyl diphosphate and farnesyl diphosphate, respectively, are essentially the same for the single and fused enzymes. The bifunctional enzymes show a more efficient conversion of isopentenyl diphosphate to epi-aristolochene than the corresponding amount of single enzymes
synthesis
-
expression system in Escherichia coli, highest expression level after 3 h induction with low concentration of IPTG and incubation at 27°C. Epiaristolochene synthase protein constitutes up to 35% of total Escherichia coli proteins
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REF.
AUTHORS
TITLE
JOURNAL
VOL.
PAGES
YEAR
ORGANISM (UNIPROT)
PUBMED ID
SOURCE
O'Maille, P.E.; Chappell, J.; Noel, J.P.
Biosynthetic potential of sesquiterpene synthase: alternative product of tobacco 5-epi-aristolochene synthase
Arch. Biochem. Biophys.
448
73-82
2006
Nicotiana tabacum
brenda
Wu, S.; Schoenbeck, M.A.; Greenhagen, B.T.; Takahashi, S.; Lee, S.; Coates, R.M.; Chappell, J.
Surrogate splicing for functional analysis of sesquiterpene synthase genes
Plant Physiol.
138
1322-1333
2005
Nicotiana tabacum (Q7X9A3)
brenda
Faraldos, J.A.; OMaille, P.E.; Dellas, N.; Noel, J.P.; Coates, R.M.
Bisabolyl-derived sesquiterpenes from tobacco 5-epi-aristolochene synthase-catalyzed cyclization of (2Z,6E)-farnesyl diphosphate
J. Am. Chem. Soc.
132
4281-4289
2010
Nicotiana tabacum
brenda
Back, K.; Yin, S.; Chappell, J.
Expression of a plant sesquiterpene cyclase gene in Escherichia coli
Arch. Biochem. Biophys.
315
527-532
1994
Nicotiana tabacum
brenda
Brodelius, M.; Lundgren, A.; Mercke, P.; Brodelius, P.
Fusion of farnesyldiphosphate synthase and epi-aristolochene synthase, a sesquiterpene cyclase involved in capsidiol biosynthesis in Nicotiana tabacum
Eur. J. Biochem.
269
3570-3577
2002
Nicotiana tabacum
brenda
Rising, K.; Starks, C.; Noel, J.; Chappell, J.
Demonstration of germacrene A as an intermediate in 5-epi-aristolochene synthase catalysis
J. Am. Chem. Soc.
122
1861-1866
2000
Nicotiana tabacum (Q40577)
-
brenda
Bohlmann, J.; Stauber, E.; Krock, B.; Oldham, N.; Gershenzon, J.; Baldwin, I.
Gene expression of 5-epi-aristolochene synthase and formation of capsidiol in roots of Nicotiana attenuata and N. sylvestris
Phytochemistry
60
109-116
2002
Nicotiana attenuata (Q84LF0), Nicotiana attenuata (Q84LF1), Nicotiana attenuata (Q84LF2)
brenda
Lee, H.; Lee, D.; Ha, S.; Jang, S.; Lee, I.; Ryu, S.; Back, K.
The characterization of transgenic rice plants expressing a pepper 5-epi aristolochene synthase, the first committed step enzyme for capsidiol synthesis in isoprenoid pathway
Plant Sci.
166
881-887
2004
Capsicum annuum
brenda
Starks, C.; Back, K.; Chappell, J.; Noel, J.
Structural basis for cyclic terpene biosynthesis by tobacco 5-epi-aristolochene synthase
Science
277
1815-1820
1997
Nicotiana tabacum (Q40577)
brenda
Diaz, J.E.; Lin, C.S.; Kunishiro, K.; Feld, B.K.; Avrantinis, S.K.; Bronson, J.; Greaves, J.; Saven, J.G.; Weiss, G.A.
Computational design and selections for an engineered, thermostable terpene synthase
Protein Sci.
20
1597-1606
2011
Nicotiana tabacum (Q40577)
brenda
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