GDO1_PSEAC
ID GDO1_PSEAC Reviewed; 347 AA.
AC Q9S3U6;
DT 07-APR-2021, integrated into UniProtKB/Swiss-Prot.
DT 01-MAY-2000, sequence version 1.
DT 03-AUG-2022, entry version 56.
DE RecName: Full=Gentisate 1,2 dioxygenase 1 {ECO:0000303|PubMed:10049846};
DE Short=GDOI {ECO:0000303|PubMed:12909360};
DE EC=1.13.11.4 {ECO:0000269|PubMed:10049846, ECO:0000269|PubMed:16237038};
GN Name=xlnE {ECO:0000303|PubMed:12909360};
OS Pseudomonas alcaligenes.
OC Bacteria; Proteobacteria; Gammaproteobacteria; Pseudomonadales;
OC Pseudomonadaceae; Pseudomonas.
OX NCBI_TaxID=43263;
RN [1]
RP NUCLEOTIDE SEQUENCE [GENOMIC DNA], INDUCTION, AND DISRUPTION PHENOTYPE.
RC STRAIN=NCIMB 9867 / P25X;
RX PubMed=12909360; DOI=10.1016/s0378-1119(03)00619-x;
RA Yeo C.C., Wong M.V.-M., Feng Y., Song K.P., Poh C.L.;
RT "Molecular characterization of an inducible gentisate 1,2-dioxygenase gene,
RT xlnE, from Pseudomonas alcaligenes NCIMB 9867.";
RL Gene 312:239-248(2003).
RN [2]
RP PROTEIN SEQUENCE OF 2-26, FUNCTION, CATALYTIC ACTIVITY, COFACTOR, ACTIVITY
RP REGULATION, BIOPHYSICOCHEMICAL PROPERTIES, AND SUBUNIT.
RC STRAIN=NCIMB 9867 / P25X;
RX PubMed=10049846; DOI=10.1128/aem.65.3.946-950.1999;
RA Feng Y., Khoo H.E., Poh C.L.;
RT "Purification and characterization of gentisate 1,2-dioxygenases from
RT Pseudomonas alcaligenes NCIB 9867 and Pseudomonas putida NCIB 9869.";
RL Appl. Environ. Microbiol. 65:946-950(1999).
RN [3]
RP FUNCTION, CATALYTIC ACTIVITY, BIOPHYSICOCHEMICAL PROPERTIES, AND
RP MUTAGENESIS OF TYR-17; VAL-36; ASN-43; SER-113; ASP-120; GLY-123; TRP-146;
RP ASN-153; GLY-164; MET-169; TYR-181; GLU-223; THR-260; VAL-284; VAL-326 AND
RP LYS-338.
RX PubMed=16237038; DOI=10.1128/jb.187.21.7543-7545.2005;
RA Tan C.L., Yeo C.C., Khoo H.E., Poh C.L.;
RT "Replacement of tyrosine 181 by phenylalanine in gentisate 1,2-dioxygenase
RT I from Pseudomonas alcaligenes NCIMB 9867 enhances catalytic activities.";
RL J. Bacteriol. 187:7543-7545(2005).
CC -!- FUNCTION: Involved in the degradation of gentisate (PubMed:10049846).
CC Catalyzes the conversion of gentisate (2,5-dihydroxybenzoate) to
CC maleylpyruvate. Exhibits broad substrate specificities towards alkyl
CC and halogenated gentisates (PubMed:10049846, PubMed:16237038).
CC {ECO:0000269|PubMed:10049846, ECO:0000269|PubMed:16237038}.
CC -!- CATALYTIC ACTIVITY:
CC Reaction=2,5-dihydroxybenzoate + O2 = 3-maleylpyruvate + H(+);
CC Xref=Rhea:RHEA:18237, ChEBI:CHEBI:15378, ChEBI:CHEBI:15379,
CC ChEBI:CHEBI:16727, ChEBI:CHEBI:58044; EC=1.13.11.4;
CC Evidence={ECO:0000269|PubMed:10049846, ECO:0000269|PubMed:16237038};
CC PhysiologicalDirection=left-to-right; Xref=Rhea:RHEA:18238;
CC Evidence={ECO:0000269|PubMed:10049846, ECO:0000269|PubMed:16237038};
CC -!- COFACTOR:
CC Name=Fe(2+); Xref=ChEBI:CHEBI:29033;
CC Evidence={ECO:0000305|PubMed:10049846};
CC -!- ACTIVITY REGULATION: Completely inhibited by the presence of 5 mM
CC Cu(2+). Partially inhibited with 5 mM Mn(2+), Zn(2+) or EDTA.
CC {ECO:0000269|PubMed:10049846}.
CC -!- BIOPHYSICOCHEMICAL PROPERTIES:
CC Kinetic parameters:
CC KM=92 uM for gentisate {ECO:0000269|PubMed:10049846};
CC KM=86.02 uM for gentisate {ECO:0000269|PubMed:16237038};
CC KM=76 uM for 3-methylgentisate {ECO:0000269|PubMed:10049846};
CC KM=62.34 uM for 3-methylgentisate {ECO:0000269|PubMed:16237038};
CC KM=15 uM for 3-bromogentisate {ECO:0000269|PubMed:10049846};
CC KM=45.91 uM for 3-bromogentisate {ECO:0000269|PubMed:16237038};
CC KM=53 uM for 3-isopropylgentisate {ECO:0000269|PubMed:10049846};
CC KM=79.37 uM for 3-isopropylgentisate {ECO:0000269|PubMed:16237038};
CC KM=76.69 uM for 3-fluorogentisate {ECO:0000269|PubMed:16237038};
CC KM=23.26 uM for 4-chlorogentisate {ECO:0000269|PubMed:16237038};
CC KM=74.26 uM for 4-methylgentisate {ECO:0000269|PubMed:16237038};
CC Note=kcat is 2433 min(-1) with gentisate as substrate. kcat is 3098
CC min(-1) with 3-methylgentisate as substrate. kcat is 613 min(-1) with
CC 3-bromogentisate as substrate. kcat is 1573 min(-1) with 3-
CC isopropylgentisate as substrate. {ECO:0000269|PubMed:10049846};
CC pH dependence:
CC Optimum pH is around 8.0. {ECO:0000269|PubMed:10049846};
CC Temperature dependence:
CC Optimum temperature is 50 degrees Celsius.
CC {ECO:0000269|PubMed:10049846};
CC -!- SUBUNIT: Homotetramer. {ECO:0000269|PubMed:10049846}.
CC -!- INDUCTION: Shows relatively high basal levels of expression under non-
CC inducing conditions. Induced by aromatic substrates, especially 3-
CC hydroxybenzoate. {ECO:0000269|PubMed:12909360}.
CC -!- DISRUPTION PHENOTYPE: Knockout mutant can still grow on minimal agar
CC plates containing gentisate as the sole carbon source but GDO activity
CC cannot be detected in uninduced cells. {ECO:0000269|PubMed:12909360}.
CC -!- SIMILARITY: Belongs to the gentisate 1,2-dioxygenase family.
CC {ECO:0000305}.
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DR EMBL; AF173167; AAD49427.1; -; Genomic_DNA.
DR AlphaFoldDB; Q9S3U6; -.
DR SMR; Q9S3U6; -.
DR BioCyc; MetaCyc:MON-18566; -.
DR SABIO-RK; Q9S3U6; -.
DR GO; GO:0047922; F:gentisate 1,2-dioxygenase activity; IEA:UniProtKB-EC.
DR GO; GO:0019439; P:aromatic compound catabolic process; IEA:UniProtKB-KW.
DR Gene3D; 2.60.120.10; -; 1.
DR InterPro; IPR013096; Cupin_2.
DR InterPro; IPR011960; Gentisate_dOase.
DR InterPro; IPR014710; RmlC-like_jellyroll.
DR InterPro; IPR011051; RmlC_Cupin_sf.
DR Pfam; PF07883; Cupin_2; 1.
DR SUPFAM; SSF51182; SSF51182; 1.
DR TIGRFAMs; TIGR02272; gentisate_1_2; 1.
PE 1: Evidence at protein level;
KW Aromatic hydrocarbons catabolism; Dioxygenase; Direct protein sequencing;
KW Iron; Oxidoreductase.
FT INIT_MET 1
FT /note="Removed"
FT /evidence="ECO:0000269|PubMed:10049846"
FT CHAIN 2..347
FT /note="Gentisate 1,2 dioxygenase 1"
FT /id="PRO_0000452280"
FT MUTAGEN 17
FT /note="Y->C: 1.6-fold increase in relative activity towards
FT gentisate."
FT /evidence="ECO:0000269|PubMed:16237038"
FT MUTAGEN 36
FT /note="V->A: Loss of activity."
FT /evidence="ECO:0000269|PubMed:16237038"
FT MUTAGEN 43
FT /note="N->T: 3.2-fold increase in relative activity towards
FT gentisate."
FT /evidence="ECO:0000269|PubMed:16237038"
FT MUTAGEN 113
FT /note="S->P: Loss of activity."
FT /evidence="ECO:0000269|PubMed:16237038"
FT MUTAGEN 120
FT /note="D->K: Loss of activity."
FT /evidence="ECO:0000269|PubMed:16237038"
FT MUTAGEN 123
FT /note="G->N: 25% decrease in activity."
FT /evidence="ECO:0000269|PubMed:16237038"
FT MUTAGEN 146
FT /note="W->T: Loss of activity."
FT /evidence="ECO:0000269|PubMed:16237038"
FT MUTAGEN 153
FT /note="N->H: 1.8-fold increase in relative activity towards
FT gentisate."
FT /evidence="ECO:0000269|PubMed:16237038"
FT MUTAGEN 164
FT /note="G->T: Loss of activity."
FT /evidence="ECO:0000269|PubMed:16237038"
FT MUTAGEN 169
FT /note="M->H: Loss of activity."
FT /evidence="ECO:0000269|PubMed:16237038"
FT MUTAGEN 181
FT /note="Y->D,H: Does not improve specific activity towards
FT gentisate."
FT /evidence="ECO:0000269|PubMed:16237038"
FT MUTAGEN 181
FT /note="Y->F: 4.6-fold increase in relative activity towards
FT gentisate. Also increases relative activities towards 3-
FT methyl-, 4-methyl-, 3-bromo- and 3-fluorogentisates."
FT /evidence="ECO:0000269|PubMed:16237038"
FT MUTAGEN 223
FT /note="E->A: No change in relative activity towards
FT gentisate."
FT /evidence="ECO:0000269|PubMed:16237038"
FT MUTAGEN 260
FT /note="T->C: Loss of activity."
FT /evidence="ECO:0000269|PubMed:16237038"
FT MUTAGEN 284
FT /note="V->I: 2.6-fold increase in relative activity towards
FT gentisate."
FT /evidence="ECO:0000269|PubMed:16237038"
FT MUTAGEN 326
FT /note="V->Q: Loss of activity."
FT /evidence="ECO:0000269|PubMed:16237038"
FT MUTAGEN 338
FT /note="K->Y: 1.5-fold increase in relative activity towards
FT gentisate."
FT /evidence="ECO:0000269|PubMed:16237038"
FT CONFLICT 22
FT /note="S -> C (in Ref. 2; AA sequence)"
FT /evidence="ECO:0000305"
SQ SEQUENCE 347 AA; 39230 MW; 416AD087EE3065DF CRC64;
MSFTEKPAVT KERKEFYSKL ESHDLAPLWE VLNEVVTTKP KSNCAPHLWE FEVAKEFLME
AGTLITAKEA ERRVLILENP GLKGLSRITT SLYAGLQLIL PGEVAPTHRH SQSALRFVVD
GGGACTSVDG ERTTMQVGDF VITPPWAWHD HVNDSDKPMI WMDGLDLPMV TLFDTSFAEG
YGEDIQEITR PNGDSLARYG ANMLPVDFKQ KGLSSPIFNY PYERSREALE AMKKANEWDP
CHGLKMQYIN PLDGMAAMPT ISSFIQLLPK EFRTQTYRST DATVFSVIEG QGKTRIGDKV
FFWKAKDTFV VPSWYPVEHE ASSDAVLFSY SDRVAQQKLG FWRESRN
