ID   HKM5_ASPHA              Reviewed;         512 AA.
AC   P0DUL3;
DT   02-JUN-2021, integrated into UniProtKB/Swiss-Prot.
DT   02-JUN-2021, sequence version 1.
DT   03-AUG-2022, entry version 5.
DE   RecName: Full=Cytochrome P450 monooxygenase hkm5 {ECO:0000303|PubMed:33242032};
DE            EC=1.-.-.- {ECO:0000269|PubMed:33242032};
DE   AltName: Full=Hancockiamides biosynthesis cluster protein 5 {ECO:0000303|PubMed:33242032};
OS   Aspergillus hancockii.
OC   Eukaryota; Fungi; Dikarya; Ascomycota; Pezizomycotina; Eurotiomycetes;
OC   Eurotiomycetidae; Eurotiales; Aspergillaceae; Aspergillus.
OX   NCBI_TaxID=1873369;
RN   [1]
RP   NUCLEOTIDE SEQUENCE [LARGE SCALE GENOMIC DNA].
RC   STRAIN=FRR 3425 / CBS 142004 / DTO 360-G7;
RA   Gilchrist C.L.M., Chooi Y.H.;
RL   Submitted (APR-2019) to the EMBL/GenBank/DDBJ databases.
RN   [2]
RP   FUNCTION, CATALYTIC ACTIVITY, PATHWAY, AND BIOTECHNOLOGY.
RX   PubMed=33242032; DOI=10.1039/d0ob02243h;
RA   Li H., Lacey A.E., Shu S., Kalaitzis J.A., Vuong D., Crombie A., Hu J.,
RA   Gilchrist C.L.M., Lacey E., Piggott A.M., Chooi Y.H.;
RT   "Hancockiamides: phenylpropanoid piperazines from Aspergillus hancockii are
RT   biosynthesised by a versatile dual single-module NRPS pathway.";
RL   Org. Biomol. Chem. 19:587-595(2021).
CC   -!- FUNCTION: Cytochrome P450 monooxygenase; part of the gene cluster that
CC       mediates the biosynthesis of hancockiamides, an unusual new family of
CC       N-cinnamoylated piperazines (PubMed:33242032). The NRPS hkm10 and the
CC       NmrA-like reductase hkm9 are proposed to convert two molecules of L-Phe
CC       to the intermediary piperazine called xenocockiamide A (Probable).
CC       Xenocockiamide A is then converted to hancockiamide D via a series of
CC       hydroxylations and O-methylations (Probable). The tyrosinase hkm6 may
CC       catalyze an aromatic hydroxylation, then the 2-oxoglutarate-dependent
CC       Fe(II) dioxygenase hkm4 and the FAD-dependent phenol hydroxylase hkm7
CC       may catalyze consecutive hydroxylations to install 2 more hydroxy
CC       groups, and the methyltransferase hkm8 probably catalyzes two
CC       methylations using 2 molecules of S-adenosyl-L-methionine (SAM)
CC       (Probable). The NRPS hkm11 activates and transfers trans-cinnamate
CC       supplied by the PAL hkm12 to hancockiamide D and produces hancockiamide
CC       A (PubMed:33242032). NRPS Hkm11 has the flexibility to tolerate the
CC       bulky hancockiamide G as a substrate and the absence of the acetyl-
CC       transferase hkm3 opens up the opportunity for hkm11 to introduce a
CC       second N-cinnamoyl moiety (PubMed:33242032). The cytochrome P450
CC       monooxygenase hkm5 catalyzes the methylenedioxy bridge formation,
CC       converting hancockiamide A into hancockiamide G (PubMed:33242032). Hkm5
CC       can also convert hancockiamide B into hancockiamide C, and
CC       hancockiamide D into hancockiamide H (PubMed:33242032). The N-
CC       acetyltransferase hkm3 finally transfers an acetyl group to 1-N of
CC       piperazine, converting hancockiamide A into hancockiamide B and
CC       hancockiamide G into hancockiamide C (PubMed:33242032).
CC       {ECO:0000269|PubMed:33242032, ECO:0000305|PubMed:33242032}.
CC   -!- COFACTOR:
CC       Name=heme; Xref=ChEBI:CHEBI:30413;
CC         Evidence={ECO:0000250|UniProtKB:P04798};
CC   -!- PATHWAY: Secondary metabolite biosynthesis.
CC       {ECO:0000269|PubMed:33242032}.
CC   -!- SUBCELLULAR LOCATION: Membrane {ECO:0000255}; Single-pass membrane
CC       protein {ECO:0000255}.
CC   -!- BIOTECHNOLOGY: Hancockiamide D displays potent cytotoxic activity
CC       against murine myeloma NS-1 cells, suggesting a potential antitumour
CC       application (PubMed:33242032). More interestingly, hancockiamide C, the
CC       likely end metabolite of the hkm pathway, shows potent Arabidopsis
CC       thaliana seed anti-germination activity, but is inactive against the
CC       monocot Eragrostis tef seed, suggesting that it could be a herbicidal
CC       lead targeting monocots (PubMed:33242032). The herbicidal activity of
CC       hancockiamide C could be due to its phenylpropanoid-like structural
CC       features, which may act on the plant lignan pathways, and hence
CC       warrants further investigations (PubMed:33242032).
CC       {ECO:0000269|PubMed:33242032}.
CC   -!- SIMILARITY: Belongs to the cytochrome P450 family. {ECO:0000305}.
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DR   EMBL; MBFL02000005; KAF7597145.1; -; Genomic_DNA.
DR   AlphaFoldDB; P0DUL3; -.
DR   SMR; P0DUL3; -.
DR   GO; GO:0016021; C:integral component of membrane; IEA:UniProtKB-KW.
DR   GO; GO:0020037; F:heme binding; IEA:InterPro.
DR   GO; GO:0005506; F:iron ion binding; IEA:InterPro.
DR   GO; GO:0004497; F:monooxygenase activity; IEA:UniProtKB-KW.
DR   GO; GO:0016705; F:oxidoreductase activity, acting on paired donors, with incorporation or reduction of molecular oxygen; IEA:InterPro.
DR   Gene3D; 1.10.630.10; -; 1.
DR   InterPro; IPR001128; Cyt_P450.
DR   InterPro; IPR002401; Cyt_P450_E_grp-I.
DR   InterPro; IPR036396; Cyt_P450_sf.
DR   Pfam; PF00067; p450; 1.
DR   PRINTS; PR00463; EP450I.
DR   PRINTS; PR00385; P450.
DR   SUPFAM; SSF48264; SSF48264; 1.
PE   1: Evidence at protein level;
KW   Glycoprotein; Heme; Iron; Membrane; Metal-binding; Monooxygenase;
KW   Oxidoreductase; Transmembrane; Transmembrane helix.
FT   CHAIN           1..512
FT                   /note="Cytochrome P450 monooxygenase hkm5"
FT                   /id="PRO_0000452932"
FT   TRANSMEM        18..38
FT                   /note="Helical"
FT                   /evidence="ECO:0000255"
FT   BINDING         456
FT                   /ligand="heme"
FT                   /ligand_id="ChEBI:CHEBI:30413"
FT                   /ligand_part="Fe"
FT                   /ligand_part_id="ChEBI:CHEBI:18248"
FT                   /note="axial binding residue"
FT                   /evidence="ECO:0000250|UniProtKB:P04798"
FT   CARBOHYD        184
FT                   /note="N-linked (GlcNAc...) asparagine"
FT                   /evidence="ECO:0000255|PROSITE-ProRule:PRU00498"
FT   CARBOHYD        263
FT                   /note="N-linked (GlcNAc...) asparagine"
FT                   /evidence="ECO:0000255|PROSITE-ProRule:PRU00498"
FT   CARBOHYD        275
FT                   /note="N-linked (GlcNAc...) asparagine"
FT                   /evidence="ECO:0000255|PROSITE-ProRule:PRU00498"
FT   CARBOHYD        374
FT                   /note="N-linked (GlcNAc...) asparagine"
FT                   /evidence="ECO:0000255|PROSITE-ProRule:PRU00498"
FT   CARBOHYD        419
FT                   /note="N-linked (GlcNAc...) asparagine"
FT                   /evidence="ECO:0000255|PROSITE-ProRule:PRU00498"
SQ   SEQUENCE   512 AA;  58106 MW;  C4828DB7E8627288 CRC64;
     METPTPPLPS KDQLFPPLIQ LVRALLWVLV ITIGGAIVQR LFFHPLRKIP GPLTAAISGW
     DEFYHNIWRD GEWCKTYPKL HKEYNSPVIR IGPNHVHLND IDAYETVFRV GTNFYKDKTF
     YTCADNDGSI FSLCDRDEHS ERRKVLSSLF SKQAAEMTAP KVMSKLNELL DFMITQSKEG
     KACNITDLFR ALAINWVADT LLGDCGDVVT YAETKPDLLE DIDGLSKLIP TLRFFPYLIP
     TLNSLAPSTS PAGVAKFKKI CENYTRPRIN DPIKNISQRS RASVVELLIA HRHEVYHKPP
     TVDYLAEEAF TFIDAGVDTT GGTLVAAIYH ILRDPGILRR LREELDESQL FLSKGTPIDF
     KKLGNLPYLN AVINESHRIW PALPGPLPRV VPPEGLQVGS FFVPSGTILS STHHCLHYNE
     TVFPEPKKFK PERWLRTDKW EGDRYLNPYS RGSRACIGIN LAQMELRLTL GHLFSHYDLQ
     LCEPTLSSLE WKDHFVAHPK APVMIHIGFR KA