HOA4_PARXL
ID HOA4_PARXL Reviewed; 346 AA.
AC P51015; Q13FU0;
DT 01-OCT-1996, integrated into UniProtKB/Swiss-Prot.
DT 19-SEP-2006, sequence version 2.
DT 03-AUG-2022, entry version 115.
DE RecName: Full=4-hydroxy-2-oxovalerate aldolase 4 {ECO:0000255|HAMAP-Rule:MF_01656};
DE Short=HOA 4 {ECO:0000255|HAMAP-Rule:MF_01656};
DE EC=4.1.3.39 {ECO:0000255|HAMAP-Rule:MF_01656, ECO:0000269|PubMed:19476337};
DE AltName: Full=4-hydroxy-2-keto-pentanoic acid aldolase 4 {ECO:0000255|HAMAP-Rule:MF_01656};
DE AltName: Full=4-hydroxy-2-oxohexanoate aldolase;
DE EC=4.1.3.43 {ECO:0000269|PubMed:19476337};
DE AltName: Full=4-hydroxy-2-oxopentanoate aldolase 4 {ECO:0000255|HAMAP-Rule:MF_01656};
GN Name=bphI; OrderedLocusNames=Bxeno_C1121; ORFNames=Bxe_C1187;
OS Paraburkholderia xenovorans (strain LB400).
OC Bacteria; Proteobacteria; Betaproteobacteria; Burkholderiales;
OC Burkholderiaceae; Paraburkholderia.
OX NCBI_TaxID=266265;
RN [1]
RP NUCLEOTIDE SEQUENCE [GENOMIC DNA].
RC STRAIN=LB400;
RX PubMed=8026764; DOI=10.1016/0378-1119(94)90196-1;
RA Hofer B., Backhaus S., Timmis K.N.;
RT "The biphenyl/polychlorinated biphenyl-degradation locus (bph) of
RT Pseudomonas sp. LB400 encodes four additional metabolic enzymes.";
RL Gene 144:9-16(1994).
RN [2]
RP NUCLEOTIDE SEQUENCE [LARGE SCALE GENOMIC DNA].
RC STRAIN=LB400;
RX PubMed=17030797; DOI=10.1073/pnas.0606924103;
RA Chain P.S.G., Denef V.J., Konstantinidis K.T., Vergez L.M., Agullo L.,
RA Reyes V.L., Hauser L., Cordova M., Gomez L., Gonzalez M., Land M., Lao V.,
RA Larimer F., LiPuma J.J., Mahenthiralingam E., Malfatti S.A., Marx C.J.,
RA Parnell J.J., Ramette A., Richardson P., Seeger M., Smith D., Spilker T.,
RA Sul W.J., Tsoi T.V., Ulrich L.E., Zhulin I.B., Tiedje J.M.;
RT "Burkholderia xenovorans LB400 harbors a multi-replicon, 9.73-Mbp genome
RT shaped for versatility.";
RL Proc. Natl. Acad. Sci. U.S.A. 103:15280-15287(2006).
RN [3]
RP FUNCTION, CATALYTIC ACTIVITY, COFACTOR, SUBSTRATE SPECIFICITY,
RP BIOPHYSICOCHEMICAL PROPERTIES, SUBUNIT, AND COMPLEX WITH BPHJ.
RC STRAIN=LB400;
RX PubMed=19476337; DOI=10.1021/bi9006644;
RA Baker P., Pan D., Carere J., Rossi A., Wang W., Seah S.Y.K.;
RT "Characterization of an aldolase-dehydrogenase complex that exhibits
RT substrate channeling in the polychlorinated biphenyls degradation
RT pathway.";
RL Biochemistry 48:6551-6558(2009).
RN [4]
RP FUNCTION, SUBSTRATE SPECIFICITY, STEREOSPECIFICITY, KINETIC PARAMETERS, AND
RP KINETIC MECHANISM.
RC STRAIN=LB400;
RX PubMed=20364820; DOI=10.1021/bi100251u;
RA Wang W., Baker P., Seah S.Y.;
RT "Comparison of two metal-dependent pyruvate aldolases related by convergent
RT evolution: substrate specificity, kinetic mechanism, and substrate
RT channeling.";
RL Biochemistry 49:3774-3782(2010).
RN [5]
RP KINETIC PARAMETERS, MUTAGENESIS OF ARG-16; HIS-20; LEU-87; LEU-89 AND
RP TYR-290, ACTIVE SITES, AND CATALYTIC MECHANISM.
RC STRAIN=LB400;
RX PubMed=21425833; DOI=10.1021/bi101947g;
RA Baker P., Carere J., Seah S.Y.;
RT "Probing the molecular basis of substrate specificity, stereospecificity,
RT and catalysis in the class II pyruvate aldolase, BphI.";
RL Biochemistry 50:3559-3569(2011).
RN [6]
RP MUTAGENESIS OF HIS-20; LEU-89; TYR-290; GLY-322 AND GLY-323, AND ALDEHYDE
RP CHANNELING MECHANISM.
RC STRAIN=LB400;
RX PubMed=21838275; DOI=10.1021/bi200960j;
RA Carere J., Baker P., Seah S.Y.;
RT "Investigating the molecular determinants for substrate channeling in BphI-
RT BphJ, an aldolase-dehydrogenase complex from the polychlorinated biphenyls
RT degradation pathway.";
RL Biochemistry 50:8407-8416(2011).
RN [7]
RP MUTAGENESIS OF LEU-87 AND TYR-290.
RC STRAIN=LB400;
RX PubMed=22081904; DOI=10.1021/ja208754r;
RA Baker P., Seah S.Y.;
RT "Rational design of stereoselectivity in the class II pyruvate aldolase
RT BphI.";
RL J. Am. Chem. Soc. 134:507-513(2012).
CC -!- FUNCTION: Catalyzes the retro-aldol cleavage of both 4-hydroxy-2-
CC oxopentanoate (HOPA) and 4-hydroxy-2-oxohexanoate (HOHA) to pyruvate
CC and acetaldehyde or propanaldehyde, respectively. The aldehydes
CC produced by this reaction are directly channeled from BphI to the
CC dehydrogenase BphJ, ensuring that these toxic aldehydes are sequestered
CC from cellular components. Is involved in the meta-cleavage pathway for
CC the degradation of polychlorinated biphenyls (PCBs). Appears to be
CC stereospecific since it can cleave (4S)-4-hydroxy-2-oxopentanoate but
CC not the (4R) isomer. Also exhibits a secondary oxaloacetate
CC decarboxylase activity. Finally, is also able to catalyze the reverse
CC reaction, albeit much less efficiently, i.e. the condensation of
CC aldehyde acceptors of two to three carbons in length with pyruvate.
CC This aldol addition reaction is stereospecific; the condensation of
CC acetaldehyde and pyruvate with BphI produces only the (4S)-4-hydroxy-2-
CC oxopentanoate isomer. Aldehyde channeling in the BphI-BphJ complex can
CC occur in reverse, from the dehydrogenase to the aldolase active sites,
CC and the BphJ reductive deacylation reaction increases 4-fold when BphI
CC is catalyzing the aldol addition reaction. Therefore, the BphI-BphJ
CC enzyme complex exhibits unique bidirectionality in substrate channeling
CC and allosteric activation. {ECO:0000269|PubMed:19476337,
CC ECO:0000269|PubMed:20364820}.
CC -!- CATALYTIC ACTIVITY:
CC Reaction=(S)-4-hydroxy-2-oxopentanoate = acetaldehyde + pyruvate;
CC Xref=Rhea:RHEA:22624, ChEBI:CHEBI:15343, ChEBI:CHEBI:15361,
CC ChEBI:CHEBI:73143; EC=4.1.3.39; Evidence={ECO:0000255|HAMAP-
CC Rule:MF_01656, ECO:0000269|PubMed:19476337};
CC -!- CATALYTIC ACTIVITY:
CC Reaction=(S)-4-hydroxy-2-oxohexanoate = propanal + pyruvate;
CC Xref=Rhea:RHEA:36003, ChEBI:CHEBI:15361, ChEBI:CHEBI:17153,
CC ChEBI:CHEBI:73142; EC=4.1.3.43;
CC Evidence={ECO:0000269|PubMed:19476337};
CC -!- COFACTOR:
CC Name=Mn(2+); Xref=ChEBI:CHEBI:29035;
CC Evidence={ECO:0000269|PubMed:19476337};
CC Name=Cd(2+); Xref=ChEBI:CHEBI:48775;
CC Evidence={ECO:0000269|PubMed:19476337};
CC Name=Co(2+); Xref=ChEBI:CHEBI:48828;
CC Evidence={ECO:0000269|PubMed:19476337};
CC Note=Divalent metal cation. Has the highest activity with Mn(2+) as
CC cofactor. Can also use Cd(2+) at low concentrations (0.01-0.1 mM) or
CC Co(2+), although with less efficiency. Mg(2+) and Ni(2+) are very poor
CC metal cofactors. {ECO:0000269|PubMed:19476337};
CC -!- ACTIVITY REGULATION: Competitively inhibited by oxalate. Also inhibited
CC by high concentrations of Cd(2+) (1 mM) in vitro. Appears to be
CC allosterically activated by aldehyde turnover occurring in BphJ, partly
CC via NADH.
CC -!- BIOPHYSICOCHEMICAL PROPERTIES:
CC Kinetic parameters:
CC KM=89 uM for (4S)-4-hydroxy-2-oxopentanoate (in the presence of NADH
CC at pH 8 and 25 degrees Celsius) {ECO:0000269|PubMed:19476337,
CC ECO:0000269|PubMed:20364820, ECO:0000269|PubMed:21425833};
CC KM=0.22 mM for racemic 4-hydroxy-2-oxopentanoate (in the presence of
CC NADH at pH 8 and 25 degrees Celsius) {ECO:0000269|PubMed:19476337,
CC ECO:0000269|PubMed:20364820, ECO:0000269|PubMed:21425833};
CC KM=1.12 mM for racemic 4-hydroxy-2-oxopentanoate (in the absence of
CC NADH at pH 8 and 25 degrees Celsius) {ECO:0000269|PubMed:19476337,
CC ECO:0000269|PubMed:20364820, ECO:0000269|PubMed:21425833};
CC KM=0.18 mM for racemic 4-hydroxy-2-oxohexanoate (in the presence of
CC NADH at pH 8 and 25 degrees Celsius) {ECO:0000269|PubMed:19476337,
CC ECO:0000269|PubMed:20364820, ECO:0000269|PubMed:21425833};
CC KM=0.35 mM for racemic 4-hydroxy-2-oxoheptanoate (in the presence of
CC NADH at pH 8 and 25 degrees Celsius) {ECO:0000269|PubMed:19476337,
CC ECO:0000269|PubMed:20364820, ECO:0000269|PubMed:21425833};
CC KM=64.28 mM for acetaldehyde (in the absence of NADH at pH 8 and 25
CC degrees Celsius) {ECO:0000269|PubMed:19476337,
CC ECO:0000269|PubMed:20364820, ECO:0000269|PubMed:21425833};
CC KM=135.9 mM for propanaldehyde (in the absence of NADH at pH 8 and 25
CC degrees Celsius) {ECO:0000269|PubMed:19476337,
CC ECO:0000269|PubMed:20364820, ECO:0000269|PubMed:21425833};
CC KM=13.0 mM for pyruvate (at pH 8 and 25 degrees Celsius)
CC {ECO:0000269|PubMed:19476337, ECO:0000269|PubMed:20364820,
CC ECO:0000269|PubMed:21425833};
CC Note=The catalytic efficiency is similar when using 4-hydroxy-2-
CC oxopentanoate or 4-hydroxy-2-oxohexanoate as substrate, but is 10-
CC fold lower with 4-hydroxy-2-oxoheptanoate. It is also 25-fold higher
CC when NADH is present than the value obtained without nucleotides.
CC Moreover, the catalytic efficiency is similar when using acetaldehyde
CC or propanaldehyde as substrate in the aldol addition reaction.;
CC pH dependence:
CC Activity increases from pH 6.5 to 9. {ECO:0000269|PubMed:19476337};
CC -!- PATHWAY: Xenobiotic degradation; polychlorinated biphenyl degradation.
CC -!- SUBUNIT: Heterotetramer composed of two BphI (aldolase) and two BphJ
CC (dehydrogenase). {ECO:0000269|PubMed:19476337}.
CC -!- MISCELLANEOUS: The aldol addition reaction proceeds via a compulsory
CC order mechanism, with pyruvate binding first.
CC -!- SIMILARITY: Belongs to the 4-hydroxy-2-oxovalerate aldolase family.
CC {ECO:0000255|HAMAP-Rule:MF_01656, ECO:0000305}.
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DR EMBL; X76500; CAA54036.1; -; Genomic_DNA.
DR EMBL; CP000272; ABE37049.1; -; Genomic_DNA.
DR RefSeq; WP_003450974.1; NZ_CP008761.1.
DR AlphaFoldDB; P51015; -.
DR SMR; P51015; -.
DR STRING; 266265.Bxe_C1187; -.
DR EnsemblBacteria; ABE37049; ABE37049; Bxe_C1187.
DR KEGG; bxb:DR64_8618; -.
DR KEGG; bxe:Bxe_C1187; -.
DR eggNOG; COG0119; Bacteria.
DR OMA; YVGGQED; -.
DR OrthoDB; 840579at2; -.
DR BRENDA; 4.1.3.39; 9987.
DR BRENDA; 4.1.3.43; 7691.
DR SABIO-RK; P51015; -.
DR UniPathway; UPA01002; -.
DR Proteomes; UP000001817; Chromosome 3.
DR GO; GO:0008701; F:4-hydroxy-2-oxovalerate aldolase activity; IEA:UniProtKB-UniRule.
DR GO; GO:0030145; F:manganese ion binding; IEA:UniProtKB-UniRule.
DR GO; GO:0019439; P:aromatic compound catabolic process; IEA:UniProtKB-UniRule.
DR CDD; cd07943; DRE_TIM_HOA; 1.
DR Gene3D; 3.20.20.70; -; 1.
DR HAMAP; MF_01656; HOA; 1.
DR InterPro; IPR017629; 4OH_2_O-val_aldolase.
DR InterPro; IPR013785; Aldolase_TIM.
DR InterPro; IPR012425; DmpG_comm.
DR InterPro; IPR035685; DRE_TIM_HOA.
DR InterPro; IPR000891; PYR_CT.
DR Pfam; PF07836; DmpG_comm; 1.
DR Pfam; PF00682; HMGL-like; 1.
DR TIGRFAMs; TIGR03217; 4OH_2_O_val_ald; 1.
DR PROSITE; PS50991; PYR_CT; 1.
PE 1: Evidence at protein level;
KW Allosteric enzyme; Aromatic hydrocarbons catabolism; Cadmium; Cobalt;
KW Lyase; Manganese; Metal-binding; Reference proteome.
FT CHAIN 1..346
FT /note="4-hydroxy-2-oxovalerate aldolase 4"
FT /id="PRO_0000064978"
FT DOMAIN 8..260
FT /note="Pyruvate carboxyltransferase"
FT /evidence="ECO:0000255|HAMAP-Rule:MF_01656"
FT ACT_SITE 20
FT /note="Proton acceptor"
FT /evidence="ECO:0000255|HAMAP-Rule:MF_01656"
FT BINDING 16..17
FT /ligand="substrate"
FT /evidence="ECO:0000255|HAMAP-Rule:MF_01656"
FT BINDING 17
FT /ligand="Mn(2+)"
FT /ligand_id="ChEBI:CHEBI:29035"
FT /evidence="ECO:0000255|HAMAP-Rule:MF_01656"
FT BINDING 170
FT /ligand="substrate"
FT /evidence="ECO:0000255|HAMAP-Rule:MF_01656"
FT BINDING 199
FT /ligand="Mn(2+)"
FT /ligand_id="ChEBI:CHEBI:29035"
FT /evidence="ECO:0000255|HAMAP-Rule:MF_01656"
FT BINDING 199
FT /ligand="substrate"
FT /evidence="ECO:0000255|HAMAP-Rule:MF_01656"
FT BINDING 201
FT /ligand="Mn(2+)"
FT /ligand_id="ChEBI:CHEBI:29035"
FT /evidence="ECO:0000255|HAMAP-Rule:MF_01656"
FT BINDING 290
FT /ligand="substrate"
FT /evidence="ECO:0000255|HAMAP-Rule:MF_01656"
FT SITE 16
FT /note="Transition state stabilizer"
FT /evidence="ECO:0000255|HAMAP-Rule:MF_01656"
FT SITE 87
FT /note="Important for aldehyde specificity, and governs
FT stereochemical control"
FT SITE 89
FT /note="Important for aldehyde specificity; governs
FT substrate alkyl chain length"
FT SITE 290
FT /note="Governs stereochemical control"
FT MUTAGEN 16
FT /note="R->A: Loss of aldol cleavage activity."
FT /evidence="ECO:0000269|PubMed:21425833"
FT MUTAGEN 16
FT /note="R->K: 4000-fold decrease in the catalytic efficiency
FT of the aldol cleavage reaction."
FT /evidence="ECO:0000269|PubMed:21425833"
FT MUTAGEN 20
FT /note="H->A,S: 100-fold decrease in the catalytic
FT efficiency of the aldol cleavage reaction. Dramatic
FT reduction in acetaldehyde and propanaldehyde channeling
FT efficiency by more than 70%."
FT /evidence="ECO:0000269|PubMed:21425833,
FT ECO:0000269|PubMed:21838275"
FT MUTAGEN 87
FT /note="L->A: 32-fold reduction in the catalytic efficiency
FT with acetaldehyde as substrate of the aldol addition
FT reaction, but no change in the catalytic efficiency using
FT propanaldehyde; thus, exhibits a 40-fold preference for
FT propanaldehyde over acetaldehyde."
FT /evidence="ECO:0000269|PubMed:21425833,
FT ECO:0000269|PubMed:22081904"
FT MUTAGEN 87
FT /note="L->N,W: Loss of aldolase activity (with either
FT enantiomer of HOPA), but retains some decarboxylase
FT activity for the smaller oxaloacetate substrate. In the
FT retro-aldol cleavage reaction, is inactive toward 4(S)-HOPA
FT but is active toward 4(R)-HOPA, albeit with a great
FT reduction in catalytic efficiency, and in the aldol
FT addition reaction, produces also exclusively the 4(R)-
FT enantiomer; when associated with F-290."
FT /evidence="ECO:0000269|PubMed:21425833,
FT ECO:0000269|PubMed:22081904"
FT MUTAGEN 89
FT /note="L->A: As the wild-type enzyme, exhibits similar
FT catalytic efficiency with acetaldehyde or propanaldehyde as
FT substrate in the aldol addition reaction but displays
FT higher catalytic efficiency with longer aldehydes (50-fold
FT increase using pentaldehyde). Shows a reduction in aldehyde
FT channeling efficiency by 30%."
FT /evidence="ECO:0000269|PubMed:21425833,
FT ECO:0000269|PubMed:21838275"
FT MUTAGEN 290
FT /note="Y->F: Loss of stereochemical control as the mutant
FT is able to catalyze the aldol cleavage of substrates with
FT both R and S configurations at C4 with similar kinetic
FT parameters. 3.5-fold decrease in the catalytic efficiency
FT of the aldol cleavage reaction. Reduction in aldehyde
FT channeling efficiency by more than 30%. In the retro-aldol
FT cleavage reaction, is inactive toward 4(S)-HOPA but is
FT active toward 4(R)-HOPA, albeit with a great reduction in
FT catalytic efficiency, and in the aldol addition reaction,
FT produces also exclusively the 4(R)-enantiomer; when
FT associated with N-87 or W-87."
FT /evidence="ECO:0000269|PubMed:21425833,
FT ECO:0000269|PubMed:21838275, ECO:0000269|PubMed:22081904"
FT MUTAGEN 290
FT /note="Y->S: Loss of stereochemical control as the mutant
FT is able to catalyze the aldol cleavage of substrates with
FT both R and S configurations at C4 with similar kinetic
FT parameters. 3.5-fold decrease in the catalytic efficiency
FT of the aldol cleavage reaction."
FT /evidence="ECO:0000269|PubMed:21425833,
FT ECO:0000269|PubMed:21838275, ECO:0000269|PubMed:22081904"
FT MUTAGEN 322
FT /note="G->A: Displays a reduction in aldehyde channeling
FT efficiency of about 20%."
FT /evidence="ECO:0000269|PubMed:21838275"
FT MUTAGEN 322
FT /note="G->F,L: Unable to channel either acetaldehyde or
FT propanaldehyde."
FT /evidence="ECO:0000269|PubMed:21838275"
FT MUTAGEN 323
FT /note="G->A: Able to channel butyraldehyde (with less
FT efficiency than wild-type) but not its isomer
FT isobutyraldehyde."
FT /evidence="ECO:0000269|PubMed:21838275"
FT MUTAGEN 323
FT /note="G->F: Unable to channel either acetaldehyde or
FT propanaldehyde."
FT /evidence="ECO:0000269|PubMed:21838275"
FT MUTAGEN 323
FT /note="G->L: Able to channel acetaldehyde but not the
FT larger propanaldehyde."
FT /evidence="ECO:0000269|PubMed:21838275"
FT CONFLICT 263
FT /note="A -> S (in Ref. 1; CAA54036)"
FT /evidence="ECO:0000305"
FT CONFLICT 302
FT /note="A -> E (in Ref. 1; CAA54036)"
FT /evidence="ECO:0000305"
SQ SEQUENCE 346 AA; 36798 MW; BD4A951D513C97CE CRC64;
MKLEGKKVTV HDMTLRDGMH PKRHQMTLEQ MKSIACGLDA AGIPLIEVTH GDGLGGSSVN
YGFPAHSDEE YLGAVIPLMK QAKVSALLLP GIGTVEHLKM AKDLGVNTIR VATHCTEADV
SEQHITQSRK LGLDTVGFLM MAHMASPEKL VSQALLMQGY GANCIYVTDS AGYMLPDDVK
ARLSAVRAAL KPETELGFHG HHNLAMGVAN SIAAIEAGAT RIDAAAAGLG AGAGNTPMEV
FIAVCARMGI ETGVDVFKIQ DVAEDLVVPI MDHVIRIDRD SLTLGYAGVY SSFLLFAKRA
SAKYGVPARD ILVELGRRGM VGGQEDMIED TAMTMARERG LTLTAA
