HPAH_ACIBA
ID HPAH_ACIBA Reviewed; 422 AA.
AC Q6Q272;
DT 22-FEB-2012, integrated into UniProtKB/Swiss-Prot.
DT 05-JUL-2004, sequence version 1.
DT 03-AUG-2022, entry version 80.
DE RecName: Full=p-hydroxyphenylacetate 3-hydroxylase, oxygenase component;
DE EC=1.14.14.9 {ECO:0000269|PubMed:11683878};
DE AltName: Full=4-HPA 3-hydroxylase {ECO:0000250|UniProtKB:Q57160};
DE AltName: Full=4-HPA 3-monooxygenase large component {ECO:0000250|UniProtKB:Q57160};
DE AltName: Full=p-hydroxyphenylacetate 3-hydroxylase C2 component;
GN Name=C2-hpah {ECO:0000312|EMBL:AAS75429.1};
OS Acinetobacter baumannii.
OC Bacteria; Proteobacteria; Gammaproteobacteria; Moraxellales; Moraxellaceae;
OC Acinetobacter; Acinetobacter calcoaceticus/baumannii complex.
OX NCBI_TaxID=470;
RN [1] {ECO:0000305, ECO:0000312|EMBL:AAS75429.1}
RP NUCLEOTIDE SEQUENCE [GENOMIC DNA], PROTEIN SEQUENCE OF 1-35, FUNCTION,
RP CATALYTIC ACTIVITY, AND KINETIC PARAMETERS.
RX PubMed=15451173; DOI=10.1016/j.bbaexp.2004.08.003;
RA Thotsaporn K., Sucharitakul J., Wongratana J., Suadee C., Chaiyen P.;
RT "Cloning and expression of p-hydroxyphenylacetate 3-hydroxylase from
RT Acinetobacter baumannii: evidence of the divergence of enzymes in the class
RT of two-protein component aromatic hydroxylases.";
RL Biochim. Biophys. Acta 1680:60-66(2004).
RN [2] {ECO:0000305}
RP FUNCTION, CATALYTIC ACTIVITY, ACTIVITY REGULATION, KINETIC PARAMETERS,
RP PATHWAY, AND SUBUNIT.
RX PubMed=11683878; DOI=10.1046/j.1432-1033.2001.02490.x;
RA Chaiyen P., Suadee C., Wilairat P.;
RT "A novel two-protein component flavoprotein hydroxylase.";
RL Eur. J. Biochem. 268:5550-5561(2001).
RN [3] {ECO:0000305}
RP FUNCTION, AND ACTIVITY REGULATION.
RX PubMed=16042421; DOI=10.1021/bi050615e;
RA Sucharitakul J., Chaiyen P., Entsch B., Ballou D.P.;
RT "The reductase of p-hydroxyphenylacetate 3-hydroxylase from Acinetobacter
RT baumannii requires p-hydroxyphenylacetate for effective catalysis.";
RL Biochemistry 44:10434-10442(2005).
RN [4] {ECO:0000305}
RP FUNCTION, AND CATALYTIC ACTIVITY.
RX PubMed=16627482; DOI=10.1074/jbc.m512385200;
RA Sucharitakul J., Chaiyen P., Entsch B., Ballou D.P.;
RT "Kinetic mechanisms of the oxygenase from a two-component enzyme, p-
RT hydroxyphenylacetate 3-hydroxylase from Acinetobacter baumannii.";
RL J. Biol. Chem. 281:17044-17053(2006).
RN [5] {ECO:0000305}
RP FUNCTION, AND ACTIVITY REGULATION.
RX PubMed=17595116; DOI=10.1021/bi7006614;
RA Sucharitakul J., Phongsak T., Entsch B., Svasti J., Chaiyen P.,
RA Ballou D.P.;
RT "Kinetics of a two-component p-hydroxyphenylacetate hydroxylase explain how
RT reduced flavin is transferred from the reductase to the oxygenase.";
RL Biochemistry 46:8611-8623(2007).
RN [6] {ECO:0000305}
RP FUNCTION, AND PH DEPENDENCE.
RX PubMed=21030590; DOI=10.1074/jbc.m110.163881;
RA Ruangchan N., Tongsook C., Sucharitakul J., Chaiyen P.;
RT "pH-dependent studies reveal an efficient hydroxylation mechanism of the
RT oxygenase component of p-hydroxyphenylacetate 3-hydroxylase.";
RL J. Biol. Chem. 286:223-233(2011).
RN [7] {ECO:0000305}
RP MUTAGENESIS OF SER-171 AND HIS-396.
RX PubMed=21680741; DOI=10.1074/jbc.m111.241836;
RA Thotsaporn K., Chenprakhon P., Sucharitakul J., Mattevi A., Chaiyen P.;
RT "Stabilization of C4a-hydroperoxyflavin in a two-component flavin-dependent
RT monooxygenase is achieved through interactions at flavin N5 and C4a
RT atoms.";
RL J. Biol. Chem. 286:28170-28180(2011).
RN [8] {ECO:0000305}
RP MUTAGENESIS OF HIS-120 AND SER-146.
RX PubMed=22052902; DOI=10.1074/jbc.m111.284463;
RA Tongsook C., Sucharitakul J., Thotsaporn K., Chaiyen P.;
RT "Interactions with the substrate phenolic group are essential for
RT hydroxylation by the oxygenase component of p-hydroxyphenylacetate 3-
RT hydroxylase.";
RL J. Biol. Chem. 286:44491-44502(2011).
RN [9] {ECO:0000305, ECO:0000312|PDB:2JBS}
RP X-RAY CRYSTALLOGRAPHY (2.30 ANGSTROMS) OF APOENZYME AND IN COMPLEXES WITH
RP REDUCED FMN AND 4-HYDROXYPHENYLACETATE, AND SUBUNIT.
RX PubMed=17227849; DOI=10.1073/pnas.0608381104;
RA Alfieri A., Fersini F., Ruangchan N., Prongjit M., Chaiyen P., Mattevi A.;
RT "Structure of the monooxygenase component of a two-component flavoprotein
RT monooxygenase.";
RL Proc. Natl. Acad. Sci. U.S.A. 104:1177-1182(2007).
CC -!- FUNCTION: Oxygenase component of a two-component system that utilizes
CC reduced FMN (FMNH2) supplied by the reductase component to catalyze the
CC hydroxylation of 4-hydroxyphenylacetic acid, leading to the production
CC of 3,4-dihydroxyphenylacetate (3,4-DHPA). Also utilizes other reduced
CC flavins such as FADH2 and reduced riboflavin to a lesser extent. Only
CC the compounds with a hydroxyl group in the para (p-) position can be
CC hydroxylated. May also oxidize phenol to catechol, and hydroxylate
CC other phenol derivatives. {ECO:0000269|PubMed:11683878,
CC ECO:0000269|PubMed:15451173, ECO:0000269|PubMed:16042421,
CC ECO:0000269|PubMed:16627482, ECO:0000269|PubMed:17595116,
CC ECO:0000269|PubMed:21030590}.
CC -!- CATALYTIC ACTIVITY:
CC Reaction=4-hydroxyphenylacetate + FMNH2 + O2 = 3,4-
CC dihydroxyphenylacetate + FMN + H(+) + H2O; Xref=Rhea:RHEA:59880,
CC ChEBI:CHEBI:15377, ChEBI:CHEBI:15378, ChEBI:CHEBI:15379,
CC ChEBI:CHEBI:17612, ChEBI:CHEBI:48999, ChEBI:CHEBI:57618,
CC ChEBI:CHEBI:58210; Evidence={ECO:0000269|PubMed:11683878,
CC ECO:0000269|PubMed:15451173, ECO:0000269|PubMed:16627482};
CC -!- CATALYTIC ACTIVITY:
CC Reaction=4-hydroxyphenylacetate + FADH2 + O2 = 3,4-
CC dihydroxyphenylacetate + FAD + H(+) + H2O; Xref=Rhea:RHEA:30595,
CC ChEBI:CHEBI:15377, ChEBI:CHEBI:15378, ChEBI:CHEBI:15379,
CC ChEBI:CHEBI:17612, ChEBI:CHEBI:48999, ChEBI:CHEBI:57692,
CC ChEBI:CHEBI:58307; EC=1.14.14.9;
CC Evidence={ECO:0000269|PubMed:11683878, ECO:0000269|PubMed:15451173,
CC ECO:0000269|PubMed:16627482};
CC -!- ACTIVITY REGULATION: Inhibited by flavin concentrations greater than 15
CC uM. Also inhibited by excess p-hydroxyphenylacetate (HPA).
CC {ECO:0000269|PubMed:11683878, ECO:0000269|PubMed:16042421,
CC ECO:0000269|PubMed:17595116}.
CC -!- BIOPHYSICOCHEMICAL PROPERTIES:
CC Kinetic parameters:
CC KM=19 uM for 4-hydroxyphenylacetate (with FMN as cosubstrate)
CC {ECO:0000269|PubMed:11683878};
CC KM=14 uM for 4-hydroxyphenylacetate (with FAD as cosubstrate)
CC {ECO:0000269|PubMed:11683878};
CC KM=25 uM for 4-hydroxyphenylacetate (with FMN as cosubstrate)
CC {ECO:0000269|PubMed:15451173};
CC KM=15 uM for 4-hydroxyphenylacetate (with FAD as cosubstrate)
CC {ECO:0000269|PubMed:15451173};
CC Note=Km values measured using the C1-C2 complex.;
CC pH dependence:
CC Optimum pH is 6-10. {ECO:0000269|PubMed:21030590};
CC -!- PATHWAY: Aromatic compound metabolism; 4-hydroxyphenylacetate
CC degradation; pyruvate and succinate semialdehyde from 4-
CC hydroxyphenylacetate: step 1/7. {ECO:0000269|PubMed:11683878}.
CC -!- SUBUNIT: Homotetramer. The p-hydroxyphenylacetate 3-hydroxylase (HpaH)
CC is composed of an oxygenase component C2 and a reductase component C1.
CC {ECO:0000269|PubMed:11683878, ECO:0000269|PubMed:17227849}.
CC -!- SIMILARITY: Belongs to the HpaH/HsaA monooxygenase family.
CC {ECO:0000305}.
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DR EMBL; AY566612; AAS75429.1; -; Genomic_DNA.
DR PDB; 2JBR; X-ray; 2.30 A; A/B/C/D=1-422.
DR PDB; 2JBS; X-ray; 2.80 A; A/B/C/D=1-422.
DR PDB; 2JBT; X-ray; 2.80 A; A/B/C/D=1-422.
DR PDBsum; 2JBR; -.
DR PDBsum; 2JBS; -.
DR PDBsum; 2JBT; -.
DR AlphaFoldDB; Q6Q272; -.
DR SMR; Q6Q272; -.
DR DIP; DIP-60804N; -.
DR STRING; 470.IX87_20440; -.
DR PRIDE; Q6Q272; -.
DR PATRIC; fig|470.1293.peg.2565; -.
DR eggNOG; COG1960; Bacteria.
DR BRENDA; 1.14.14.9; 98.
DR UniPathway; UPA00208; UER00416.
DR EvolutionaryTrace; Q6Q272; -.
DR GO; GO:0052881; F:4-hydroxyphenylacetate 3-monooxygenase activity; IEA:UniProtKB-EC.
DR GO; GO:0050660; F:flavin adenine dinucleotide binding; IEA:InterPro.
DR GO; GO:0016627; F:oxidoreductase activity, acting on the CH-CH group of donors; IEA:InterPro.
DR GO; GO:0019439; P:aromatic compound catabolic process; IEA:UniProtKB-KW.
DR Gene3D; 1.10.540.10; -; 1.
DR Gene3D; 2.40.110.10; -; 1.
DR InterPro; IPR013107; Acyl-CoA_DH_C.
DR InterPro; IPR046373; Acyl-CoA_Oxase/DH_mid-dom_sf.
DR InterPro; IPR036250; AcylCo_DH-like_C.
DR InterPro; IPR037069; AcylCoA_DH/ox_N_sf.
DR InterPro; IPR009100; AcylCoA_DH/oxidase_NM_dom.
DR Pfam; PF08028; Acyl-CoA_dh_2; 1.
DR SUPFAM; SSF47203; SSF47203; 1.
DR SUPFAM; SSF56645; SSF56645; 1.
PE 1: Evidence at protein level;
KW 3D-structure; Aromatic hydrocarbons catabolism; Direct protein sequencing;
KW FAD; Flavoprotein; FMN; Monooxygenase; Nucleotide-binding; Oxidoreductase.
FT CHAIN 1..422
FT /note="p-hydroxyphenylacetate 3-hydroxylase, oxygenase
FT component"
FT /id="PRO_0000415755"
FT BINDING 112
FT /ligand="FMN"
FT /ligand_id="ChEBI:CHEBI:58210"
FT /evidence="ECO:0000269|PubMed:17227849"
FT BINDING 120
FT /ligand="substrate"
FT BINDING 146..148
FT /ligand="FMN"
FT /ligand_id="ChEBI:CHEBI:58210"
FT /evidence="ECO:0000269|PubMed:17227849"
FT BINDING 146
FT /ligand="substrate"
FT BINDING 169..171
FT /ligand="FMN"
FT /ligand_id="ChEBI:CHEBI:58210"
FT /evidence="ECO:0000269|PubMed:17227849"
FT BINDING 263..266
FT /ligand="substrate"
FT BINDING 292
FT /ligand="FMN"
FT /ligand_id="ChEBI:CHEBI:58210"
FT /evidence="ECO:0000269|PubMed:17227849"
FT BINDING 296
FT /ligand="FMN"
FT /ligand_id="ChEBI:CHEBI:58210"
FT /evidence="ECO:0000269|PubMed:17227849"
FT BINDING 296
FT /ligand="substrate"
FT BINDING 374..375
FT /ligand="FMN"
FT /ligand_id="ChEBI:CHEBI:58210"
FT /evidence="ECO:0000269|PubMed:17227849"
FT BINDING 396..397
FT /ligand="FMN"
FT /ligand_id="ChEBI:CHEBI:58210"
FT /evidence="ECO:0000269|PubMed:17227849"
FT MUTAGEN 120
FT /note="H->D,N: Loss of hydroxylation activity. 7 to 10-fold
FT higher rate constant for hydrogen peroxide elimination."
FT /evidence="ECO:0000269|PubMed:22052902"
FT MUTAGEN 120
FT /note="H->K: 170-fold higher rate constant for hydrogen
FT peroxide elimination."
FT /evidence="ECO:0000269|PubMed:22052902"
FT MUTAGEN 146
FT /note="S->A: Decrease in rate constant for hydroxylation by
FT 6-fold."
FT /evidence="ECO:0000269|PubMed:22052902"
FT MUTAGEN 146
FT /note="S->C: Decrease in rate constant for hydroxylation by
FT 45-fold and decreased enzymatic efficiency at pH greater
FT than 9."
FT /evidence="ECO:0000269|PubMed:22052902"
FT MUTAGEN 171
FT /note="S->A: Failure to form reaction intermediate; when
FT associated with V-396. Decrease in rate constant for the
FT formation of intermediate by 11-fold. Increase in rate
FT constant for hydrogen peroxide elimination by 1400-fold."
FT /evidence="ECO:0000269|PubMed:21680741"
FT MUTAGEN 171
FT /note="S->T: Increase in rate constant for hydrogen
FT peroxide elimination by 8-fold."
FT /evidence="ECO:0000269|PubMed:21680741"
FT MUTAGEN 396
FT /note="H->A: Decrease in rate constant for the formation of
FT the reaction intermediate by 100-fold. Denatured above pH
FT 10."
FT /evidence="ECO:0000269|PubMed:21680741"
FT MUTAGEN 396
FT /note="H->K: Reduced binding with flavin. Lower rate
FT constant. Denatured above pH 10."
FT /evidence="ECO:0000269|PubMed:21680741"
FT MUTAGEN 396
FT /note="H->N: Decrease in rate constant for the formation of
FT the reaction intermediate by 30-fold. Denatured above pH
FT 10."
FT /evidence="ECO:0000269|PubMed:21680741"
FT MUTAGEN 396
FT /note="H->R: Reduced binding with flavin. Lower rate
FT constant. Denatured above pH 10."
FT /evidence="ECO:0000269|PubMed:21680741"
FT MUTAGEN 396
FT /note="H->V: Failure to form reaction intermediate; when
FT associated with A-171. Decrease in rate constant for the
FT formation of the reaction intermediate by 300-fold.
FT Denatured above pH 10."
FT /evidence="ECO:0000269|PubMed:21680741"
FT STRAND 27..29
FT /evidence="ECO:0007829|PDB:2JBR"
FT HELIX 40..53
FT /evidence="ECO:0007829|PDB:2JBR"
FT HELIX 55..61
FT /evidence="ECO:0007829|PDB:2JBR"
FT HELIX 66..74
FT /evidence="ECO:0007829|PDB:2JBR"
FT HELIX 77..79
FT /evidence="ECO:0007829|PDB:2JBR"
FT STRAND 80..82
FT /evidence="ECO:0007829|PDB:2JBR"
FT HELIX 84..86
FT /evidence="ECO:0007829|PDB:2JBR"
FT HELIX 93..106
FT /evidence="ECO:0007829|PDB:2JBR"
FT HELIX 108..124
FT /evidence="ECO:0007829|PDB:2JBR"
FT HELIX 129..136
FT /evidence="ECO:0007829|PDB:2JBR"
FT STRAND 144..147
FT /evidence="ECO:0007829|PDB:2JBR"
FT STRAND 152..157
FT /evidence="ECO:0007829|PDB:2JBR"
FT STRAND 160..170
FT /evidence="ECO:0007829|PDB:2JBR"
FT HELIX 173..175
FT /evidence="ECO:0007829|PDB:2JBR"
FT STRAND 177..186
FT /evidence="ECO:0007829|PDB:2JBR"
FT STRAND 192..200
FT /evidence="ECO:0007829|PDB:2JBR"
FT HELIX 201..203
FT /evidence="ECO:0007829|PDB:2JBR"
FT STRAND 205..207
FT /evidence="ECO:0007829|PDB:2JBR"
FT STRAND 212..215
FT /evidence="ECO:0007829|PDB:2JBR"
FT HELIX 216..218
FT /evidence="ECO:0007829|PDB:2JBR"
FT STRAND 221..231
FT /evidence="ECO:0007829|PDB:2JBR"
FT HELIX 232..234
FT /evidence="ECO:0007829|PDB:2JBR"
FT STRAND 235..237
FT /evidence="ECO:0007829|PDB:2JBR"
FT HELIX 238..242
FT /evidence="ECO:0007829|PDB:2JBR"
FT TURN 247..250
FT /evidence="ECO:0007829|PDB:2JBR"
FT STRAND 256..260
FT /evidence="ECO:0007829|PDB:2JBR"
FT HELIX 262..266
FT /evidence="ECO:0007829|PDB:2JBR"
FT HELIX 269..289
FT /evidence="ECO:0007829|PDB:2JBR"
FT TURN 295..297
FT /evidence="ECO:0007829|PDB:2JBR"
FT HELIX 305..337
FT /evidence="ECO:0007829|PDB:2JBR"
FT HELIX 344..369
FT /evidence="ECO:0007829|PDB:2JBR"
FT HELIX 374..377
FT /evidence="ECO:0007829|PDB:2JBR"
FT HELIX 382..393
FT /evidence="ECO:0007829|PDB:2JBR"
FT TURN 397..399
FT /evidence="ECO:0007829|PDB:2JBR"
FT HELIX 401..412
FT /evidence="ECO:0007829|PDB:2JBR"
SQ SEQUENCE 422 AA; 46950 MW; 01BBA6D3C5CA5F80 CRC64;
MENTVLNLDS DVIHACEAIF QPIRLVYTHA QTPDVSGVSM LEKIQQILPQ IAKNAESAEQ
LRRVPDENIK LLKEIGLHRA FQPKVYGGLE MSLPDFANCI VTLAGACAGT AWAFSLLCTH
SHQIAMFSKQ LQDEIWLKDP DATASSSIAP FGKVEEVEGG IILNGDYGWS SGCDHAEYAI
VGFNRFDADG NKIYSFGVIP RSDYEIVDNW YAQAIKSSGS KMLKLVNVFI PEYRISKAKD
MMEGKSAGFG LYPDSKIFYT PYRPYFASGF SAVSLGIAER MIEAFKEKQR NRVRAYTGAN
VGLATPALMR IAESTHQVAA ARALLEKTWE DHRIHGLNHQ YPNKETLAFW RTNQAYAVKM
CIEAVDRLMA AAGATSFMDN SELQRLFRDA HMTGAHAYTD YDVCAQILGR ELMGMEPDPT
MV
