ID   PRX2_PENRO              Reviewed;         342 AA.
AC   Q03471;
DT   01-OCT-1993, integrated into UniProtKB/Swiss-Prot.
DT   01-OCT-1993, sequence version 1.
DT   27-MAR-2024, entry version 119.
DE   RecName: Full=Aristolochene synthase {ECO:0000303|PubMed:15186158};
DE            Short=AS {ECO:0000303|PubMed:15186158};
DE            EC=4.2.3.9 {ECO:0000269|PubMed:15186158, ECO:0000269|PubMed:8440737};
DE   AltName: Full=PR-toxin biosynthesis protein 2 {ECO:0000303|PubMed:24239699};
DE   AltName: Full=Sesquiterpene cyclase {ECO:0000303|PubMed:15186158};
GN   Name=prx2 {ECO:0000303|PubMed:24239699};
GN   Synonyms=ari1 {ECO:0000303|PubMed:15186158}, ORF2
GN   {ECO:0000303|PubMed:27921136};
OS   Penicillium roqueforti.
OC   Eukaryota; Fungi; Dikarya; Ascomycota; Pezizomycotina; Eurotiomycetes;
OC   Eurotiomycetidae; Eurotiales; Aspergillaceae; Penicillium.
OX   NCBI_TaxID=5082;
RN   [1]
RP   NUCLEOTIDE SEQUENCE [GENOMIC DNA], PROTEIN SEQUENCE OF 56-70; 76-84;
RP   245-251 AND 323-336, FUNCTION, AND CATALYTIC ACTIVITY.
RC   STRAIN=ATCC 10110 / BCRC 32359 / CBS 221.30 / JCM 22842 / NBRC 5459 /
RC   NCTC 588 / NRRL 849;
RX   PubMed=8440737; DOI=10.1016/S0021-9258(18)53644-9;
RA   Proctor R.H., Hohn T.M.;
RT   "Aristolochene synthase. Isolation, characterization, and bacterial
RT   expression of a sesquiterpenoid biosynthetic gene (Ari1) from Penicillium
RT   roqueforti.";
RL   J. Biol. Chem. 268:4543-4548(1993).
RN   [2]
RP   FUNCTION.
RX   PubMed=16345540; DOI=10.1128/aem.39.4.770-776.1980;
RA   Moreau S., Lablache-Combier A., Biguet J.;
RT   "Production of eremofortins A, B, and C relative to formation of PR toxin
RT   by Penicillium roqueforti.";
RL   Appl. Environ. Microbiol. 39:770-776(1980).
RN   [3]
RP   FUNCTION, CATALYTIC ACTIVITY, BIOPHYSICOCHEMICAL PROPERTIES, AND
RP   MUTAGENESIS OF TYR-92; ASP-115; ASN-244; SER-248 AND GLU-252.
RX   PubMed=15186158; DOI=10.1021/ja0499593;
RA   Felicetti B., Cane D.E.;
RT   "Aristolochene synthase: mechanistic analysis of active site residues by
RT   site-directed mutagenesis.";
RL   J. Am. Chem. Soc. 126:7212-7221(2004).
RN   [4]
RP   FUNCTION, AND DISRUPTION PHENOTYPE.
RX   PubMed=24239699; DOI=10.1016/j.fgb.2013.10.009;
RA   Hidalgo P.I., Ullan R.V., Albillos S.M., Montero O., Fernandez-Bodega M.A.,
RA   Garcia-Estrada C., Fernandez-Aguado M., Martin J.F.;
RT   "Molecular characterization of the PR-toxin gene cluster in Penicillium
RT   roqueforti and Penicillium chrysogenum: cross talk of secondary metabolite
RT   pathways.";
RL   Fungal Genet. Biol. 62:11-24(2014).
RN   [5]
RP   FUNCTION.
RX   PubMed=26274339; DOI=10.1002/anie.201506128;
RA   Riclea R., Dickschat J.S.;
RT   "Identification of intermediates in the biosynthesis of PR toxin by
RT   Penicillium roqueforti.";
RL   Angew. Chem. Int. Ed. 54:12167-12170(2015).
RN   [6]
RP   FUNCTION, AND PATHWAY.
RX   PubMed=27921136; DOI=10.1007/s00253-016-7995-5;
RA   Hidalgo P.I., Poirier E., Ullan R.V., Piqueras J., Meslet-Cladiere L.,
RA   Coton E., Coton M.;
RT   "Penicillium roqueforti PR toxin gene cluster characterization.";
RL   Appl. Microbiol. Biotechnol. 101:2043-2056(2017).
RN   [7]
RP   X-RAY CRYSTALLOGRAPHY (2.5 ANGSTROMS) IN COMPLEX WITH SUBSTRATE.
RX   PubMed=10825154; DOI=10.1074/jbc.m000433200;
RA   Caruthers J.M., Kang I., Rynkiewicz M.J., Cane D.E., Christianson D.W.;
RT   "Crystal structure determination of aristolochene synthase from the blue
RT   cheese mold, Penicillium roqueforti.";
RL   J. Biol. Chem. 275:25533-25539(2000).
CC   -!- FUNCTION: Aristolochene synthase; part of the gene cluster that
CC       mediates the biosynthesis of PR-toxin, a bicyclic sesquiterpene
CC       belonging to the eremophilane class and acting as a mycotoxin
CC       (PubMed:24239699, PubMed:27921136). The first step of the pathway is
CC       catalyzed by the aristolochene synthase which performs the cyclization
CC       of trans,trans-farnesyl diphosphate (FPP) to the bicyclic sesquiterpene
CC       aristolochene (PubMed:8440737, PubMed:15186158, PubMed:24239699).
CC       Following the formation of aristolochene, the non-oxygenated
CC       aristolochene is converted to the trioxygenated intermediate
CC       eremofortin B, via 7-epi-neopetasone (PubMed:24239699,
CC       PubMed:26274339). This conversion appears to involve three enzymes, a
CC       hydroxysterol oxidase-like enzyme, the quinone-oxidase prx3 that forms
CC       the quinone-type-structure in the bicyclic nucleus of aristolochene
CC       with the C8-oxo group and the C-3 hydroxyl group, and the P450
CC       monooxygenase ORF6 that introduces the epoxide at the double bond
CC       between carbons 1 and 2 (PubMed:24239699, PubMed:27921136). No monoxy
CC       or dioxy-intermediates have been reported to be released to the broth,
CC       so these three early oxidative reactions may be coupled together
CC       (PubMed:24239699). Eremofortin B is further oxidized by another P450
CC       monooxygenase, that introduces a second epoxide between carbons 7 and
CC       11 prior to acetylation to eremofortin A by the acetyltransferase ORF8
CC       (PubMed:16345540, PubMed:24239699, PubMed:27921136). The second
CC       epoxidation may be performed by a second P450 monooxygenase
CC       (PubMed:24239699). After the acetylation step, the conversion of
CC       eremofortin A to eremofortin C and then to PR-toxin requires only two
CC       enzymes (PubMed:24239699). First the conversion of eremofortin A to
CC       eremofortin C proceeds by oxidation of the side chain of the molecule
CC       at C-12 and is catalyzed by the short-chain oxidoreductase prx1
CC       (PubMed:16345540, PubMed:24239699). The cytochrome P450 monooxygenase
CC       ORF5 also plays a role in this step (PubMed:27921136). The primary
CC       alcohol formed at C-12 is finally oxidized by the short-chain alcohol
CC       dehydrogenase prx4 that forms PR-toxin (PubMed:16345540,
CC       PubMed:24239699). {ECO:0000269|PubMed:15186158,
CC       ECO:0000269|PubMed:16345540, ECO:0000269|PubMed:24239699,
CC       ECO:0000269|PubMed:26274339, ECO:0000269|PubMed:27921136,
CC       ECO:0000269|PubMed:8440737}.
CC   -!- CATALYTIC ACTIVITY:
CC       Reaction=(2E,6E)-farnesyl diphosphate = (+)-aristolochene +
CC         diphosphate; Xref=Rhea:RHEA:19825, ChEBI:CHEBI:33019,
CC         ChEBI:CHEBI:43445, ChEBI:CHEBI:175763; EC=4.2.3.9;
CC         Evidence={ECO:0000269|PubMed:15186158, ECO:0000269|PubMed:8440737};
CC   -!- COFACTOR:
CC       Name=Mg(2+); Xref=ChEBI:CHEBI:18420;
CC         Evidence={ECO:0000250|UniProtKB:Q9UR08};
CC       Note=Binds 3 Mg(2+) ions per subunit. {ECO:0000250|UniProtKB:Q9UR08};
CC   -!- BIOPHYSICOCHEMICAL PROPERTIES:
CC       Kinetic parameters:
CC         KM=0.6 uM for (2E,6E)-farnesyl diphosphate
CC         {ECO:0000269|PubMed:15186158};
CC   -!- PATHWAY: Sesquiterpene biosynthesis; aristolochene biosynthesis;
CC       aristolochene from farnesyl diphosphate: step 1/1.
CC       {ECO:0000269|PubMed:15186158, ECO:0000269|PubMed:8440737}.
CC   -!- SUBUNIT: Homodimer. {ECO:0000250|UniProtKB:Q9UR08}.
CC   -!- DISRUPTION PHENOTYPE: Reduces the production of PR-toxin and leads to a
CC       large increase in mycophenolic acid production (PubMed:24239699).
CC       {ECO:0000269|PubMed:24239699}.
CC   -!- SIMILARITY: Belongs to the terpene synthase family. {ECO:0000305}.
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DR   EMBL; L05193; AAA33694.1; -; Genomic_DNA.
DR   PIR; A45462; A45462.
DR   PDB; 1DGP; X-ray; 2.80 A; A/B=40-339.
DR   PDB; 1DI1; X-ray; 2.50 A; A/B=40-339.
DR   PDBsum; 1DGP; -.
DR   PDBsum; 1DI1; -.
DR   AlphaFoldDB; Q03471; -.
DR   SMR; Q03471; -.
DR   KEGG; ag:AAA33694; -.
DR   OMA; FQMSGNE; -.
DR   OrthoDB; 2431512at2759; -.
DR   PhylomeDB; Q03471; -.
DR   BioCyc; MetaCyc:MONOMER-16547; -.
DR   BRENDA; 4.2.3.9; 4638.
DR   SABIO-RK; Q03471; -.
DR   UniPathway; UPA00177; UER00582.
DR   EvolutionaryTrace; Q03471; -.
DR   GO; GO:0045483; F:aristolochene synthase activity; IEA:UniProtKB-EC.
DR   GO; GO:0046872; F:metal ion binding; IEA:UniProtKB-KW.
DR   CDD; cd00687; Terpene_cyclase_nonplant_C1; 1.
DR   Gene3D; 1.10.600.10; Farnesyl Diphosphate Synthase; 1.
DR   InterPro; IPR008949; Isoprenoid_synthase_dom_sf.
DR   PANTHER; PTHR35201; TERPENE SYNTHASE; 1.
DR   PANTHER; PTHR35201:SF4; TERPENE SYNTHASE; 1.
DR   Pfam; PF19086; Terpene_syn_C_2; 1.
DR   SUPFAM; SSF48576; Terpenoid synthases; 1.
PE   1: Evidence at protein level;
KW   3D-structure; Direct protein sequencing; Lyase; Magnesium; Metal-binding.
FT   CHAIN           1..342
FT                   /note="Aristolochene synthase"
FT                   /id="PRO_0000064677"
FT   BINDING         115
FT                   /ligand="Mg(2+)"
FT                   /ligand_id="ChEBI:CHEBI:18420"
FT                   /ligand_label="1"
FT                   /evidence="ECO:0000250|UniProtKB:Q9UR08"
FT   BINDING         115
FT                   /ligand="Mg(2+)"
FT                   /ligand_id="ChEBI:CHEBI:18420"
FT                   /ligand_label="2"
FT                   /evidence="ECO:0000250|UniProtKB:Q9UR08"
FT   BINDING         115
FT                   /ligand="substrate"
FT                   /evidence="ECO:0000269|PubMed:10825154,
FT                   ECO:0007744|PDB:1DGP"
FT   BINDING         200
FT                   /ligand="substrate"
FT                   /evidence="ECO:0000250|UniProtKB:Q9UR08"
FT   BINDING         244
FT                   /ligand="Mg(2+)"
FT                   /ligand_id="ChEBI:CHEBI:18420"
FT                   /ligand_label="3"
FT                   /evidence="ECO:0000250|UniProtKB:Q9UR08"
FT   BINDING         244
FT                   /ligand="substrate"
FT                   /evidence="ECO:0000269|PubMed:10825154,
FT                   ECO:0007744|PDB:1DGP"
FT   BINDING         248
FT                   /ligand="Mg(2+)"
FT                   /ligand_id="ChEBI:CHEBI:18420"
FT                   /ligand_label="3"
FT                   /evidence="ECO:0000250|UniProtKB:Q9UR08"
FT   BINDING         251
FT                   /ligand="substrate"
FT                   /evidence="ECO:0000250|UniProtKB:Q9UR08"
FT   BINDING         252
FT                   /ligand="Mg(2+)"
FT                   /ligand_id="ChEBI:CHEBI:18420"
FT                   /ligand_label="3"
FT                   /evidence="ECO:0000250|UniProtKB:Q9UR08"
FT   SITE            92
FT                   /note="Important for catalytic activity"
FT                   /evidence="ECO:0000269|PubMed:10825154,
FT                   ECO:0007744|PDB:1DI1"
FT   SITE            112
FT                   /note="Important for catalytic activity"
FT                   /evidence="ECO:0000269|PubMed:10825154,
FT                   ECO:0007744|PDB:1DI1"
FT   SITE            178
FT                   /note="Important for catalytic activity"
FT                   /evidence="ECO:0000269|PubMed:10825154,
FT                   ECO:0007744|PDB:1DI1"
FT   SITE            334
FT                   /note="Important for catalytic activity"
FT                   /evidence="ECO:0000269|PubMed:10825154,
FT                   ECO:0007744|PDB:1DI1"
FT   MUTAGEN         92
FT                   /note="Y->F: Causes 100-fold reduction in kcat but a
FT                   50-fold decrease in KM, resulting in a 2-fold decrease in
FT                   catalytic efficiency."
FT                   /evidence="ECO:0000269|PubMed:15186158"
FT   MUTAGEN         115
FT                   /note="D->N: Abolishes catalytic activity."
FT                   /evidence="ECO:0000269|PubMed:15186158"
FT   MUTAGEN         244
FT                   /note="N->L: Abolishes catalytic activity."
FT                   /evidence="ECO:0000269|PubMed:15186158"
FT   MUTAGEN         248
FT                   /note="S->A: Abolishes catalytic activity; when associated
FT                   with D-252."
FT                   /evidence="ECO:0000269|PubMed:15186158"
FT   MUTAGEN         252
FT                   /note="E->D: Abolishes catalytic activity; when associated
FT                   with A-248."
FT                   /evidence="ECO:0000269|PubMed:15186158"
FT   HELIX           54..68
FT                   /evidence="ECO:0007829|PDB:1DI1"
FT   HELIX           74..83
FT                   /evidence="ECO:0007829|PDB:1DI1"
FT   HELIX           85..92
FT                   /evidence="ECO:0007829|PDB:1DI1"
FT   TURN            98..100
FT                   /evidence="ECO:0007829|PDB:1DI1"
FT   HELIX           101..120
FT                   /evidence="ECO:0007829|PDB:1DI1"
FT   HELIX           123..137
FT                   /evidence="ECO:0007829|PDB:1DI1"
FT   HELIX           148..180
FT                   /evidence="ECO:0007829|PDB:1DI1"
FT   TURN            181..183
FT                   /evidence="ECO:0007829|PDB:1DI1"
FT   HELIX           193..201
FT                   /evidence="ECO:0007829|PDB:1DI1"
FT   HELIX           204..217
FT                   /evidence="ECO:0007829|PDB:1DI1"
FT   HELIX           223..227
FT                   /evidence="ECO:0007829|PDB:1DI1"
FT   HELIX           230..248
FT                   /evidence="ECO:0007829|PDB:1DI1"
FT   TURN            249..252
FT                   /evidence="ECO:0007829|PDB:1DI1"
FT   HELIX           269..277
FT                   /evidence="ECO:0007829|PDB:1DI1"
FT   HELIX           281..307
FT                   /evidence="ECO:0007829|PDB:1DI1"
FT   HELIX           315..336
FT                   /evidence="ECO:0007829|PDB:1DI1"
SQ   SEQUENCE   342 AA;  39192 MW;  ACFBD63F4FF9BB03 CRC64;
     MATSTETISS LAQPFVHLEN PINSPLVKET IRPRNDTTIT PPPTQWSYLC HPRVKEVQDE
     VDGYFLENWK FPSFKAVRTF LDAKFSEVTC LYFPLALDDR IHFACRLLTV LFLIDDVLEH
     MSFADGEAYN NRLIPISRGD VLPDRTKPEE FILYDLWESM RAHDAELANE VLEPTFVFMR
     AQTDRARLSI HELGHYLEYR EKDVGKALLS ALMRFSMGLR LSADELQDMK ALEANCAKQL
     SVVNDIYSYD KEEEASRTGH KEGAFLCSAV KVLAEESKLG IPATKRVLWS MTREWETVHD
     EIVAEKIASP DGCSEAAKAY MKGLEYQMSG NEQWSKTTRR YN
//