METF_ECOLI
ID METF_ECOLI Reviewed; 296 AA.
AC P0AEZ1; P00394; Q2M8N7;
DT 21-JUL-1986, integrated into UniProtKB/Swiss-Prot.
DT 21-JUL-1986, sequence version 1.
DT 03-AUG-2022, entry version 130.
DE RecName: Full=5,10-methylenetetrahydrofolate reductase;
DE EC=1.5.1.20;
GN Name=metF; OrderedLocusNames=b3941, JW3913;
OS Escherichia coli (strain K12).
OC Bacteria; Proteobacteria; Gammaproteobacteria; Enterobacterales;
OC Enterobacteriaceae; Escherichia.
OX NCBI_TaxID=83333;
RN [1]
RP NUCLEOTIDE SEQUENCE [GENOMIC DNA].
RC STRAIN=K12;
RX PubMed=6356036; DOI=10.1093/nar/11.19.6723;
RA Saint-Girons I., Duchange N., Zakin M.M., Park I., Margarita D.,
RA Ferrara P., Cohen G.N.;
RT "Nucleotide sequence of metF, the E. coli structural gene for 5-10
RT methylene tetrahydrofolate reductase and of its control region.";
RL Nucleic Acids Res. 11:6723-6732(1983).
RN [2]
RP NUCLEOTIDE SEQUENCE [LARGE SCALE GENOMIC DNA].
RC STRAIN=K12 / MG1655 / ATCC 47076;
RX PubMed=8346018; DOI=10.1093/nar/21.15.3391;
RA Plunkett G. III, Burland V., Daniels D.L., Blattner F.R.;
RT "Analysis of the Escherichia coli genome. III. DNA sequence of the region
RT from 87.2 to 89.2 minutes.";
RL Nucleic Acids Res. 21:3391-3398(1993).
RN [3]
RP NUCLEOTIDE SEQUENCE [LARGE SCALE GENOMIC DNA].
RC STRAIN=K12 / MG1655 / ATCC 47076;
RX PubMed=9278503; DOI=10.1126/science.277.5331.1453;
RA Blattner F.R., Plunkett G. III, Bloch C.A., Perna N.T., Burland V.,
RA Riley M., Collado-Vides J., Glasner J.D., Rode C.K., Mayhew G.F.,
RA Gregor J., Davis N.W., Kirkpatrick H.A., Goeden M.A., Rose D.J., Mau B.,
RA Shao Y.;
RT "The complete genome sequence of Escherichia coli K-12.";
RL Science 277:1453-1462(1997).
RN [4]
RP NUCLEOTIDE SEQUENCE [LARGE SCALE GENOMIC DNA].
RC STRAIN=K12 / W3110 / ATCC 27325 / DSM 5911;
RX PubMed=16738553; DOI=10.1038/msb4100049;
RA Hayashi K., Morooka N., Yamamoto Y., Fujita K., Isono K., Choi S.,
RA Ohtsubo E., Baba T., Wanner B.L., Mori H., Horiuchi T.;
RT "Highly accurate genome sequences of Escherichia coli K-12 strains MG1655
RT and W3110.";
RL Mol. Syst. Biol. 2:E1-E5(2006).
RN [5]
RP INDUCTION, FUNCTION, CATALYTIC ACTIVITY, BIOPHYSICOCHEMICAL PROPERTIES, AND
RP COFACTOR.
RX PubMed=14275142;
RA Katzen H.M., Buchanan J.M.;
RT "Enzymatic synthesis of the methyl group of methionine. 8. Repression-
RT derepression, purification, and properties of 5,10-
RT methylenetetrahydrofolate reductase from Escherichia coli.";
RL J. Biol. Chem. 240:825-835(1965).
RN [6]
RP IDENTIFICATION.
RX PubMed=6219268; DOI=10.1007/bf00384390;
RA Zakin M.M., Greene R.C., Dautry-Varsat A., Duchange N., Ferrara P.,
RA Py M.C., Margarita D., Cohen G.N.;
RT "Construction and physical mapping of plasmids containing the metJBLF gene
RT cluster of E. coli K12.";
RL Mol. Gen. Genet. 187:101-106(1982).
RN [7]
RP CATALYTIC ACTIVITY, COFACTOR, ACTIVE SITE, AND MUTAGENESIS OF GLU-28 AND
RP ASP-120.
RX PubMed=11371182; DOI=10.1021/bi002790v;
RA Trimmer E.E., Ballou D.P., Ludwig M.L., Matthews R.G.;
RT "Folate activation and catalysis in methylenetetrahydrofolate reductase
RT from Escherichia coli: roles for aspartate 120 and glutamate 28.";
RL Biochemistry 40:6216-6226(2001).
RN [8]
RP X-RAY CRYSTALLOGRAPHY (2.5 ANGSTROMS) OF 21-294 IN COMPLEX WITH FAD,
RP BIOPHYSICOCHEMICAL PROPERTIES, COFACTOR, SUBUNIT, AND MUTAGENESIS OF
RP ALA-177.
RX PubMed=10201405; DOI=10.1038/7594;
RA Guenther B.D., Sheppard C.A., Tran P., Rozen R., Matthews R.G.,
RA Ludwig M.L.;
RT "The structure and properties of methylenetetrahydrofolate reductase from
RT Escherichia coli suggest how folate ameliorates human
RT hyperhomocysteinemia.";
RL Nat. Struct. Biol. 6:359-365(1999).
RN [9]
RP X-RAY CRYSTALLOGRAPHY (1.85 ANGSTROMS) OF WILD TYPE AND MUTANT GLN-28 IN
RP COMPLEXES WITH 5-METHYL-5,6,7,8-TETRAHYDROFOLATE; FAD AND NAD, COFACTOR,
RP SUBUNIT, MUTAGENESIS OF GLU-28, AND ACTIVE SITE.
RX PubMed=16114881; DOI=10.1021/bi050533q;
RA Pejchal R., Sargeant R., Ludwig M.L.;
RT "Structures of NADH and CH3-H4folate complexes of Escherichia coli
RT methylenetetrahydrofolate reductase reveal a spartan strategy for a ping-
RT pong reaction.";
RL Biochemistry 44:11447-11457(2005).
RN [10]
RP X-RAY CRYSTALLOGRAPHY (2.05 ANGSTROMS) OF MUTANT VAL-177 IN COMPLEX WITH
RP SUBSTRATE ANALOG LY309887 AND FAD, COFACTOR, AND SUBUNIT.
RX PubMed=16605249; DOI=10.1021/bi052294c;
RA Pejchal R., Campbell E., Guenther B.D., Lennon B.W., Matthews R.G.,
RA Ludwig M.L.;
RT "Structural perturbations in the Ala --> Val polymorphism of
RT methylenetetrahydrofolate reductase: how binding of folates may protect
RT against inactivation.";
RL Biochemistry 45:4808-4818(2006).
RN [11]
RP X-RAY CRYSTALLOGRAPHY (1.65 ANGSTROMS) ON MUTANTS GLN-28 AND LEU-223 IN
RP COMPLEXES WITH 5-METHYL-5,6,7,8-TETRAHYDROFOLATE AND FAD, CATALYTIC
RP ACTIVITY, MUTAGENESIS OF PHE-223, AND COFACTOR.
RX PubMed=19610625; DOI=10.1021/bi9007325;
RA Lee M.N., Takawira D., Nikolova A.P., Ballou D.P., Furtado V.C.,
RA Phung N.L., Still B.R., Thorstad M.K., Tanner J.J., Trimmer E.E.;
RT "Functional role for the conformationally mobile phenylalanine 223 in the
RT reaction of methylenetetrahydrofolate reductase from Escherichia coli.";
RL Biochemistry 48:7673-7685(2009).
CC -!- FUNCTION: Methylenetetrahydrofolate reductase required to generate the
CC methyl groups necessary for methionine synthetase to convert
CC homocysteine to methionine. {ECO:0000269|PubMed:14275142}.
CC -!- CATALYTIC ACTIVITY:
CC Reaction=(6S)-5-methyl-5,6,7,8-tetrahydrofolate + NADP(+) = (6R)-5,10-
CC methylene-5,6,7,8-tetrahydrofolate + H(+) + NADPH;
CC Xref=Rhea:RHEA:19817, ChEBI:CHEBI:15378, ChEBI:CHEBI:15636,
CC ChEBI:CHEBI:18608, ChEBI:CHEBI:57783, ChEBI:CHEBI:58349; EC=1.5.1.20;
CC Evidence={ECO:0000269|PubMed:11371182, ECO:0000269|PubMed:14275142,
CC ECO:0000269|PubMed:19610625};
CC -!- CATALYTIC ACTIVITY:
CC Reaction=(6S)-5-methyl-5,6,7,8-tetrahydrofolate + NAD(+) = (6R)-5,10-
CC methylene-5,6,7,8-tetrahydrofolate + H(+) + NADH;
CC Xref=Rhea:RHEA:19821, ChEBI:CHEBI:15378, ChEBI:CHEBI:15636,
CC ChEBI:CHEBI:18608, ChEBI:CHEBI:57540, ChEBI:CHEBI:57945; EC=1.5.1.20;
CC Evidence={ECO:0000269|PubMed:11371182, ECO:0000269|PubMed:14275142,
CC ECO:0000269|PubMed:19610625};
CC -!- COFACTOR:
CC Name=FAD; Xref=ChEBI:CHEBI:57692;
CC Evidence={ECO:0000269|PubMed:10201405, ECO:0000269|PubMed:11371182,
CC ECO:0000269|PubMed:14275142, ECO:0000269|PubMed:16114881,
CC ECO:0000269|PubMed:16605249, ECO:0000269|PubMed:19610625};
CC -!- BIOPHYSICOCHEMICAL PROPERTIES:
CC Kinetic parameters:
CC KM=17 uM for NADPH {ECO:0000269|PubMed:10201405,
CC ECO:0000269|PubMed:14275142};
CC KM=3.9 uM for 5,10-methylenetetrahydrofolate
CC {ECO:0000269|PubMed:10201405, ECO:0000269|PubMed:14275142};
CC pH dependence:
CC Optimum pH is 6.3-6.4. {ECO:0000269|PubMed:10201405,
CC ECO:0000269|PubMed:14275142};
CC -!- PATHWAY: One-carbon metabolism; tetrahydrofolate interconversion.
CC -!- SUBUNIT: Homotetramer. {ECO:0000269|PubMed:10201405,
CC ECO:0000269|PubMed:16114881, ECO:0000269|PubMed:16605249}.
CC -!- INDUCTION: Repressed by methionine. {ECO:0000269|PubMed:14275142}.
CC -!- SIMILARITY: Belongs to the methylenetetrahydrofolate reductase family.
CC {ECO:0000305}.
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DR EMBL; V01502; CAA24747.1; -; Genomic_DNA.
DR EMBL; L19201; AAB03073.1; -; Genomic_DNA.
DR EMBL; U00096; AAC76923.1; -; Genomic_DNA.
DR EMBL; AP009048; BAE77369.1; -; Genomic_DNA.
DR PIR; A00462; RDECMH.
DR RefSeq; NP_418376.1; NC_000913.3.
DR RefSeq; WP_000007523.1; NZ_STEB01000037.1.
DR PDB; 1B5T; X-ray; 2.50 A; A/B/C=21-294.
DR PDB; 1ZP3; X-ray; 1.85 A; A/B/C=1-296.
DR PDB; 1ZP4; X-ray; 1.85 A; A/B/C=1-296.
DR PDB; 1ZPT; X-ray; 1.95 A; A/B/C=1-296.
DR PDB; 1ZRQ; X-ray; 2.20 A; A/B/C=1-296.
DR PDB; 2FMN; X-ray; 2.05 A; A/B/C=1-296.
DR PDB; 2FMO; X-ray; 2.25 A; A/B/C=1-296.
DR PDB; 3FST; X-ray; 1.65 A; A/C/E=1-296.
DR PDB; 3FSU; X-ray; 1.70 A; A/C/E=1-296.
DR PDB; 6PEY; X-ray; 2.88 A; A/B/C=1-296.
DR PDBsum; 1B5T; -.
DR PDBsum; 1ZP3; -.
DR PDBsum; 1ZP4; -.
DR PDBsum; 1ZPT; -.
DR PDBsum; 1ZRQ; -.
DR PDBsum; 2FMN; -.
DR PDBsum; 2FMO; -.
DR PDBsum; 3FST; -.
DR PDBsum; 3FSU; -.
DR PDBsum; 6PEY; -.
DR AlphaFoldDB; P0AEZ1; -.
DR SMR; P0AEZ1; -.
DR BioGRID; 4263063; 24.
DR DIP; DIP-6848N; -.
DR IntAct; P0AEZ1; 4.
DR STRING; 511145.b3941; -.
DR DrugBank; DB03147; Flavin adenine dinucleotide.
DR PaxDb; P0AEZ1; -.
DR PRIDE; P0AEZ1; -.
DR EnsemblBacteria; AAC76923; AAC76923; b3941.
DR EnsemblBacteria; BAE77369; BAE77369; BAE77369.
DR GeneID; 66672148; -.
DR GeneID; 948432; -.
DR KEGG; ecj:JW3913; -.
DR KEGG; eco:b3941; -.
DR PATRIC; fig|1411691.4.peg.2763; -.
DR EchoBASE; EB0580; -.
DR eggNOG; COG0685; Bacteria.
DR HOGENOM; CLU_025841_0_0_6; -.
DR InParanoid; P0AEZ1; -.
DR OMA; EMHPQAR; -.
DR PhylomeDB; P0AEZ1; -.
DR BioCyc; EcoCyc:METHYLENETHFREDUCT-MON; -.
DR BioCyc; MetaCyc:METHYLENETHFREDUCT-MON; -.
DR BRENDA; 1.5.1.20; 2026.
DR BRENDA; 1.5.1.54; 2026.
DR SABIO-RK; P0AEZ1; -.
DR UniPathway; UPA00193; -.
DR EvolutionaryTrace; P0AEZ1; -.
DR PRO; PR:P0AEZ1; -.
DR Proteomes; UP000000318; Chromosome.
DR Proteomes; UP000000625; Chromosome.
DR GO; GO:0005829; C:cytosol; IBA:GO_Central.
DR GO; GO:0032991; C:protein-containing complex; IDA:EcoCyc.
DR GO; GO:0071949; F:FAD binding; IDA:EcoCyc.
DR GO; GO:0004489; F:methylenetetrahydrofolate reductase (NAD(P)H) activity; IDA:EcoCyc.
DR GO; GO:0106312; F:methylenetetrahydrofolate reductase NADH activity; IEA:RHEA.
DR GO; GO:0106313; F:methylenetetrahydrofolate reductase NADPH activity; IEA:RHEA.
DR GO; GO:0009086; P:methionine biosynthetic process; IBA:GO_Central.
DR GO; GO:0035999; P:tetrahydrofolate interconversion; IBA:GO_Central.
DR CDD; cd00537; MTHFR; 1.
DR InterPro; IPR029041; FAD-linked_oxidoreductase-like.
DR InterPro; IPR003171; Mehydrof_redctse-like.
DR InterPro; IPR004620; MTHF_reductase_bac.
DR Pfam; PF02219; MTHFR; 1.
DR SUPFAM; SSF51730; SSF51730; 1.
DR TIGRFAMs; TIGR00676; fadh2; 1.
PE 1: Evidence at protein level;
KW 3D-structure; Amino-acid biosynthesis; FAD; Flavoprotein;
KW Methionine biosynthesis; NAD; NADP; Oxidoreductase; Reference proteome.
FT CHAIN 1..296
FT /note="5,10-methylenetetrahydrofolate reductase"
FT /id="PRO_0000190261"
FT ACT_SITE 28
FT /note="Proton donor/acceptor"
FT /evidence="ECO:0000269|PubMed:11371182,
FT ECO:0000269|PubMed:16114881"
FT BINDING 28..33
FT /ligand="NAD(+)"
FT /ligand_id="ChEBI:CHEBI:57540"
FT BINDING 59..62
FT /ligand="FAD"
FT /ligand_id="ChEBI:CHEBI:57692"
FT /evidence="ECO:0000269|PubMed:10201405,
FT ECO:0000269|PubMed:16605249"
FT BINDING 59..60
FT /ligand="NAD(+)"
FT /ligand_id="ChEBI:CHEBI:57540"
FT BINDING 88
FT /ligand="FAD"
FT /ligand_id="ChEBI:CHEBI:57692"
FT /evidence="ECO:0000269|PubMed:10201405,
FT ECO:0000269|PubMed:16605249"
FT BINDING 118..120
FT /ligand="FAD"
FT /ligand_id="ChEBI:CHEBI:57692"
FT /evidence="ECO:0000269|PubMed:10201405,
FT ECO:0000269|PubMed:16605249"
FT BINDING 120
FT /ligand="substrate"
FT BINDING 131..132
FT /ligand="FAD"
FT /ligand_id="ChEBI:CHEBI:57692"
FT /evidence="ECO:0000269|PubMed:10201405,
FT ECO:0000269|PubMed:16605249"
FT BINDING 152
FT /ligand="FAD"
FT /ligand_id="ChEBI:CHEBI:57692"
FT /evidence="ECO:0000269|PubMed:10201405,
FT ECO:0000269|PubMed:16605249"
FT BINDING 156..159
FT /ligand="FAD"
FT /ligand_id="ChEBI:CHEBI:57692"
FT /evidence="ECO:0000269|PubMed:10201405,
FT ECO:0000269|PubMed:16605249"
FT BINDING 165..172
FT /ligand="FAD"
FT /ligand_id="ChEBI:CHEBI:57692"
FT /evidence="ECO:0000269|PubMed:10201405,
FT ECO:0000269|PubMed:16605249"
FT BINDING 183
FT /ligand="substrate"
FT BINDING 219
FT /ligand="substrate"
FT BINDING 223
FT /ligand="substrate"
FT BINDING 275
FT /ligand="substrate"
FT BINDING 279
FT /ligand="substrate"
FT MUTAGEN 28
FT /note="E->Q: Abolishes enzyme activity."
FT /evidence="ECO:0000269|PubMed:11371182,
FT ECO:0000269|PubMed:16114881"
FT MUTAGEN 120
FT /note="D->N: Strongly reduces enzyme activity. Strongly
FT reduces affinity for 5-methyltetrahydrofolate."
FT /evidence="ECO:0000269|PubMed:11371182"
FT MUTAGEN 177
FT /note="A->V: Increases the propensity to lose its essential
FT flavin cofactor."
FT /evidence="ECO:0000269|PubMed:10201405"
FT MUTAGEN 223
FT /note="F->A: Strongly decreases substrate and NADH
FT binding."
FT /evidence="ECO:0000269|PubMed:19610625"
FT MUTAGEN 223
FT /note="F->L: Slightly reduced enzyme activity."
FT /evidence="ECO:0000269|PubMed:19610625"
FT HELIX 4..17
FT /evidence="ECO:0007829|PDB:3FST"
FT TURN 18..21
FT /evidence="ECO:0007829|PDB:3FST"
FT STRAND 24..29
FT /evidence="ECO:0007829|PDB:3FST"
FT HELIX 35..49
FT /evidence="ECO:0007829|PDB:3FST"
FT STRAND 54..58
FT /evidence="ECO:0007829|PDB:3FST"
FT HELIX 67..81
FT /evidence="ECO:0007829|PDB:3FST"
FT STRAND 85..91
FT /evidence="ECO:0007829|PDB:3FST"
FT HELIX 96..108
FT /evidence="ECO:0007829|PDB:3FST"
FT STRAND 113..117
FT /evidence="ECO:0007829|PDB:3FST"
FT STRAND 123..125
FT /evidence="ECO:0007829|PDB:1B5T"
FT HELIX 132..142
FT /evidence="ECO:0007829|PDB:3FST"
FT STRAND 146..151
FT /evidence="ECO:0007829|PDB:3FST"
FT HELIX 162..175
FT /evidence="ECO:0007829|PDB:3FST"
FT STRAND 179..182
FT /evidence="ECO:0007829|PDB:3FST"
FT HELIX 188..200
FT /evidence="ECO:0007829|PDB:3FST"
FT STRAND 207..211
FT /evidence="ECO:0007829|PDB:1B5T"
FT HELIX 217..226
FT /evidence="ECO:0007829|PDB:3FST"
FT HELIX 233..239
FT /evidence="ECO:0007829|PDB:3FST"
FT HELIX 246..266
FT /evidence="ECO:0007829|PDB:3FST"
FT STRAND 271..275
FT /evidence="ECO:0007829|PDB:3FST"
FT HELIX 281..289
FT /evidence="ECO:0007829|PDB:3FST"
SQ SEQUENCE 296 AA; 33103 MW; B702702D2BE9521E CRC64;
MSFFHASQRD ALNQSLAEVQ GQINVSFEFF PPRTSEMEQT LWNSIDRLSS LKPKFVSVTY
GANSGERDRT HSIIKGIKDR TGLEAAPHLT CIDATPDELR TIARDYWNNG IRHIVALRGD
LPPGSGKPEM YASDLVTLLK EVADFDISVA AYPEVHPEAK SAQADLLNLK RKVDAGANRA
ITQFFFDVES YLRFRDRCVS AGIDVEIIPG ILPVSNFKQA KKFADMTNVR IPAWMAQMFD
GLDDDAETRK LVGANIAMDM VKILSREGVK DFHFYTLNRA EMSYAICHTL GVRPGL
