ID   HKM4_ASPHA              Reviewed;         305 AA.
AC   P0DUL2;
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=Dioxygenase hkm4 {ECO:0000303|PubMed:33242032};
DE            EC=1.14.11.- {ECO:0000305|PubMed:33242032};
DE   AltName: Full=Hancockiamides biosynthesis cluster protein 4 {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, 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: Dioxygenase; part of the gene cluster that mediates the
CC       biosynthesis of hancockiamides, an unusual new family of N-
CC       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=Fe cation; Xref=ChEBI:CHEBI:24875;
CC         Evidence={ECO:0000250|UniProtKB:G8GV69};
CC   -!- PATHWAY: Secondary metabolite biosynthesis.
CC       {ECO:0000305|PubMed:33242032}.
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 PhyH family. {ECO:0000305}.
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DR   EMBL; MBFL02000005; KAF7597146.1; -; Genomic_DNA.
DR   AlphaFoldDB; P0DUL2; -.
DR   SMR; P0DUL2; -.
DR   GO; GO:0051213; F:dioxygenase activity; IEA:UniProtKB-KW.
DR   GO; GO:0046872; F:metal ion binding; IEA:UniProtKB-KW.
DR   InterPro; IPR008775; Phytyl_CoA_dOase.
DR   Pfam; PF05721; PhyH; 1.
PE   1: Evidence at protein level;
KW   Dioxygenase; Iron; Metal-binding; Oxidoreductase.
FT   CHAIN           1..305
FT                   /note="Dioxygenase hkm4"
FT                   /id="PRO_0000452931"
FT   BINDING         140
FT                   /ligand="Fe cation"
FT                   /ligand_id="ChEBI:CHEBI:24875"
FT                   /evidence="ECO:0000250|UniProtKB:G8GV69"
FT   BINDING         142
FT                   /ligand="Fe cation"
FT                   /ligand_id="ChEBI:CHEBI:24875"
FT                   /evidence="ECO:0000250|UniProtKB:G8GV69"
FT   BINDING         216
FT                   /ligand="Fe cation"
FT                   /ligand_id="ChEBI:CHEBI:24875"
FT                   /evidence="ECO:0000250|UniProtKB:G8GV69"
SQ   SEQUENCE   305 AA;  33741 MW;  5A325E2619D2D326 CRC64;
     MSIPSGTEPQ IKRFSVTADP DTIFQAYQED GVVIIQGFLS PEQLDKFNRE VNPRLAHQRQ
     GYQPSLKARL MEGSLSALLP PQQKRVHNLA GFSKVFRHDI LNHGLMHELC RRAFAATGDY
     WLSSGAVIEN GPGTPEQGWH RDQPSYPVIQ AGPGTAEGMV NFFTALTDFT AEAGATQFMH
     GSHKVVGIPD GDPNHPMLIA EMKAGDSVLL SGKLVHRGGL NNTSDFFRRA LSLAISPCVL
     TPYESSIHLS RPLVESMTPL AQRMIAWRSA SIPPPYQIGM WTLNMNEVGE EMGLKYNQPY
     DEDEE