DB code: D00262

RLCP classification 9.1050.440000.8011 : Hydride transfer
5.10.9510.969 : Elimination
9.5010.536210.8011 : Hydride transfer
CATH domain 3.40.50.720 : Rossmann fold Catalytic domain
3.90.25.10 : UDP-galactose 4-epimerase; domain 1
E.C. 4.2.1.46
CSA
M-CSA
MACiE M0228

CATH domain Related DB codes (homologues)
3.40.50.720 : Rossmann fold S00543 S00551 S00552 S00553 S00602 S00604 S00605 S00608 S00610 S00625 S00319 S00328 S00329 S00330 S00331 S00332 D00456 D00457 D00458 S00324 S00320 S00325 S00326 S00327 D00459 S00335 S00336 S00334 T00219 S00339 D00513 D00001 D00002 D00003 D00005 D00007 D00008 D00010 D00012 D00017 D00018 D00023 D00027 D00028 D00031 D00032 D00033 D00034 D00035 D00037 D00048 D00071 D00476 D00481 D00482 D00490 D00492 D00494 D00545 D00601 D00603 D00604 D00605 D00615 D00845 D00857 D00858 M00161 M00171 M00210 T00002 T00010 T00011 T00015 T00227 T00247 T00408 T00414 D00827 D00274 D00275 M00035 T00109
3.90.25.10 : UDP-galactose 4-epimerase; domain 1 D00513 D00601 D00604 D00274 D00275

Uniprot Enzyme Name
UniprotKB Protein name Synonyms RefSeq Pfam
P27830 dTDP-glucose 4,6-dehydratase 2
EC 4.2.1.46
YP_026255.1 (Protein)
NC_000913.2 (DNA/RNA sequence)
YP_491651.1 (Protein)
NC_007779.1 (DNA/RNA sequence)
PF01370 (Epimerase)
[Graphical View]
Q9EU31 dTDP-glucose 4,6-dehydratase
EC 4.2.1.46
PF01370 (Epimerase)
[Graphical View]
P26391 dTDP-glucose 4,6-dehydratase
EC 4.2.1.46
NP_461042.1 (Protein)
NC_003197.1 (DNA/RNA sequence)
PF01370 (Epimerase)
[Graphical View]
P95780 dTDP-glucose 4,6-dehydratase
EC 4.2.1.46
NP_721810.1 (Protein)
NC_004350.2 (DNA/RNA sequence)
PF01370 (Epimerase)
[Graphical View]
Q8GIP9 dTDP-glucose 4,6-dehydratase
EC 4.2.1.46
YP_004401333.1 (Protein)
NC_015433.1 (DNA/RNA sequence)
PF01370 (Epimerase)
[Graphical View]
Q9ZGH3 dTDP-glucose 4,6-dehydratase
EC 4.2.1.46
PF01370 (Epimerase)
[Graphical View]

KEGG enzyme name
dTDP-glucose 4,6-dehydratase
thymidine diphosphoglucose oxidoreductase
TDP-glucose oxidoreductase
dTDP-glucose 4,6-hydro-lyase

UniprotKB: Accession Number Entry name Activity Subunit Subcellular location Cofactor
P27830 RFFG_ECOLI dTDP-glucose = dTDP-4-dehydro-6-deoxy-D-glucose + H(2)O. Homodimer. NAD.
Q9EU31 Q9EU31_SALCH dTDP-glucose = dTDP-4-dehydro-6-deoxy-D-glucose + H(2)O. NAD (By similarity).
P26391 RMLB_SALTY dTDP-glucose = dTDP-4-dehydro-6-deoxy-D-glucose + H(2)O. Homodimer. Binds 1 NAD ion per monomer.
P95780 RMLB_STRMU dTDP-glucose = dTDP-4-dehydro-6-deoxy-D-glucose + H(2)O. Homodimer. Binds 1 NAD ion per monomer.
Q8GIP9 Q8GIP9_STRSU dTDP-glucose = dTDP-4-dehydro-6-deoxy-D-glucose + H(2)O. NAD (By similarity).
Q9ZGH3 Q9ZGH3_9ACTO dTDP-glucose = dTDP-4-dehydro-6-deoxy-D-glucose + H(2)O. NAD (By similarity).

KEGG Pathways
Map code Pathways E.C.
MAP00520 Nucleotide sugars metabolism
MAP00521 Streptomycin biosynthesis
MAP00523 Polyketide sugar unit biosynthesis
MAP01055 Biosynthesis of vancomycin group antibiotics

Compound table
Cofactors Substrates Products Intermediates
KEGG-id C00003 C00842 C11907 C00001 I00091 I00092
E.C.
Compound NAD+ dTDP-glucose dTDP-4-dehydro-6-deoxy-D-glucose H2O dTDP-4-dehydro-glucose dTDP-6-deoxy-4-dehydro-5,6-ene-glucose
Type amide group,amine group,nucleotide amide group,carbohydrate,nucleotide amide group,carbohydrate,nucleotide H2O
ChEBI 15846
 15846
 		15700
 15700
 		16128
 16128
 		15377
 15377
PubChem 5893
 5893
 		443210
 443210
 		439292
 439292
 		22247451
 962
 22247451
 962
1bxkA01 Bound:NAD Unbound Unbound Unbound Unbound
1bxkB01 Bound:NAD Unbound Unbound Unbound Unbound
1g1aA01 Bound:NAD Unbound Unbound Unbound Unbound
1g1aB01 Bound:NAD Unbound Unbound Unbound Unbound
1g1aC01 Bound:NAD Unbound Unbound Unbound Unbound
1g1aD01 Bound:NAD Unbound Unbound Unbound Unbound
1keuA01 Bound:NAD Unbound Unbound Unbound Unbound
1keuB01 Bound:NAD Unbound Unbound Unbound Unbound
1kewA01 Bound:NAD Unbound Unbound Unbound Unbound
1kewB01 Bound:NAD Unbound Unbound Unbound Unbound
1kepA01 Bound:NAD Analogue:TDX Unbound Unbound Unbound
1kepB01 Bound:NAD Unbound Unbound Unbound Unbound
1kerA01 Bound:NAD Unbound Unbound Unbound Unbound
1kerB01 Bound:NAD Unbound Unbound Unbound Unbound
1ketA01 Bound:NAD Unbound Unbound Unbound Unbound
1ketB01 Bound:NAD Unbound Unbound Unbound Unbound
1oc2A01 Bound:NAD Unbound Unbound Unbound Unbound
1oc2B01 Bound:NAD Unbound Unbound Unbound Unbound
1r66A01 Bound:NAD Unbound Unbound Unbound Unbound
1r6dA01 Bound:NAD Unbound Unbound Unbound Unbound
1bxkA02 Unbound Unbound Unbound Unbound Unbound
1bxkB02 Unbound Unbound Unbound Unbound Unbound
1g1aA02 Unbound Unbound Unbound Unbound Unbound
1g1aB02 Unbound Unbound Unbound Unbound Unbound
1g1aC02 Unbound Unbound Unbound Unbound Unbound
1g1aD02 Unbound Unbound Unbound Unbound Unbound
1keuA02 Unbound Bound:DAU Unbound Unbound Unbound
1keuB02 Unbound Bound:DAU Unbound Unbound Unbound
1kewA02 Unbound Analogue:TYD Unbound Unbound Unbound
1kewB02 Unbound Analogue:TYD Unbound Unbound Unbound
1kepA02 Unbound Unbound Unbound Unbound Unbound
1kepB02 Unbound Analogue:TDX Unbound Unbound Unbound
1kerA02 Unbound Bound:DAU Unbound Unbound Unbound
1kerB02 Unbound Bound:DAU Unbound Unbound Unbound
1ketA02 Unbound Analogue:TYD Unbound Unbound Unbound
1ketB02 Unbound Analogue:TYD Unbound Unbound Unbound
1oc2A02 Unbound Analogue:TDX Unbound Unbound Unbound
1oc2B02 Unbound Analogue:TDX Unbound Unbound Unbound
1r66A02 Unbound Analogue:TYD Unbound Unbound Unbound
1r6dA02 Unbound Bound:DAU Unbound Unbound Unbound

Reference for Active-site residues
resource references E.C.
literature [8], [9], [12], [13] & [14]

Active-site residues
PDB Catalytic residues Cofactor-binding residues Modified residues Main-chain involved in catalysis Comment
1bxkA01 THR 134;ASP 135;GLU 136;TYR 160;LYS 164;ARG 224
1bxkB01 THR 534;ASP 535;GLU 536;TYR 560;LYS 564;ARG 624
1g1aA01 THR 133;ASP 134;GLU 135;TYR 167;LYS 171;ARG 231
1g1aB01 THR 133;ASP 134;GLU 135;TYR 167;LYS 171;ARG 231
1g1aC01 THR 133;ASP 134;GLU 135;TYR 167;LYS 171;ARG 231
1g1aD01 THR 133;ASP 134;GLU 135;TYR 167;LYS 171;ARG 231
1keuA01 THR 133;ASP 134;GLU 135;TYR 167;LYS 171;ARG 231
1keuB01 THR 133;ASP 134;GLU 135;TYR 167;LYS 171;ARG 231
1kewA01 THR 133;ASP 134;GLU 135;TYR 167;LYS 171;ARG 231
1kewB01 THR 133;ASP 134;GLU 135;TYR 167;LYS 171;ARG 231
1kepA01 THR 125;ASP 126;GLU 127;TYR 161;LYS 165;ARG 225
1kepB01 THR 125;ASP 126;GLU 127;TYR 161;LYS 165;ARG 225
1kerA01 THR 125;ASP 126;GLU 127;TYR 161;LYS 165;ARG 225
1kerB01 THR 125;ASP 126;GLU 127;TYR 161;LYS 165;ARG 225
1ketA01 THR 125;ASP 126;GLU 127;TYR 161;LYS 165;ARG 225
1ketB01 THR 125;ASP 126;GLU 127;TYR 161;LYS 165;ARG 225
1oc2A01 THR 125;ASP 126;GLU 127;TYR 161;LYS 165;ARG 225
1oc2B01 THR 125;ASP 126;GLU 127;TYR 161;LYS 165;ARG 225
1r66A01 THR 127;ASP 128;GLU 129;TYR 151;LYS 155;ARG 215
1r6dA01 THR 127;; ;TYR 151;LYS 155;ARG 215 mutant D128N, E129Q
1bxkA02
1bxkB02
1g1aA02
1g1aB02
1g1aC02
1g1aD02
1keuA02
1keuB02
1kewA02
1kewB02
1kepA02
1kepB02
1kerA02
1kerB02
1ketA02
1ketB02
1oc2A02
1oc2B02
1r66A02
1r6dA02

References for Catalytic Mechanism
References Sections No. of steps in catalysis
[7]
Scheme 1, Fig.1, Fig.5 2
[8]
Scheme 1, Scheme 2, Scheme 3 2
[9]
Scheme 1, Scheme 2, Scheme 3 2
[10]
Scheme 1, Scheme 3, Scheme 4, Scheme 5 3
[12]
Fig.1 5
[13]
Scheme 1, Scheme 2, Fig.4, Scheme 3
[14]
Fig.4, p.88 3
[15]
Scheme 1
[16]
Scheme 2, p.2217-2220
[17]
Fig.3, p.648-651

References
[1]
Resource
Comments
Medline ID
PubMed ID 7517391
Journal J Bacteriol
Year 1994
Volume 176
Pages 4144-56
Authors Stevenson G, Neal B, Liu D, Hobbs M, Packer NH, Batley M, Redmond JW, Lindquist L, Reeves P
Title Structure of the O antigen of Escherichia coli K-12 and the sequence of its rfb gene cluster.
Related PDB
Related UniProtKB
[2]
Resource
Comments FUNCTION
Medline ID 96032389
PubMed ID 7559340
Journal J Bacteriol
Year 1995
Volume 177
Pages 5539-46
Authors Marolda CL, Valvano MA
Title Genetic analysis of the dTDP-rhamnose biosynthesis region of the Escherichia coli VW187 (O7:K1) rfb gene cluster: identification of functional homologs of rfbB and rfbA in the rff cluster and correct location of the rffE gene.
Related PDB
Related UniProtKB P27830
[3]
Resource
Comments
Medline ID
PubMed ID 9011374
Journal Carbohydr Res
Year 1996
Volume 285
Pages 141-50
Authors Naundorf A, Klaffke W
Title Substrate specificity of native dTDP-D-glucose-4,6-dehydratase: chemo-enzymatic syntheses of artificial and naturally occurring deoxy sugars.
Related PDB
Related UniProtKB
[4]
Resource
Comments
Medline ID
PubMed ID 10462438
Journal Arch Biochem Biophys
Year 1999
Volume 369
Pages 30-41
Authors Essigmann B, Hespenheide BM, Kuhn LA, Benning C
Title Prediction of the active-site structure and NAD(+) binding in SQD1, a protein essential for sulfolipid biosynthesis in Arabidopsis.
Related PDB
Related UniProtKB
[5]
Resource
Comments
Medline ID
PubMed ID 10358040
Journal J Biol Chem
Year 1999
Volume 274
Pages 16933-9
Authors Yoshida Y, Nakano Y, Nezu T, Yamashita Y, Koga T
Title A novel NDP-6-deoxyhexosyl-4-ulose reductase in the pathway for the synthesis of thymidine diphosphate-D-fucose.
Related PDB
Related UniProtKB
[6]
Resource
Comments
Medline ID
PubMed ID 10666612
Journal Acta Crystallogr D Biol Crystallogr
Year 2000
Volume 56
Pages 222-5
Authors Allard ST, Giraud MF, Whitfield C, Messner P, Naismith JH
Title The purification, crystallization and structural elucidation of dTDP-D-glucose 4,6-dehydratase (RmlB), the second enzyme of the dTDP-L-rhamnose synthesis pathway from Salmonella enterica serovar typhimurium.
Related PDB
Related UniProtKB
[7]
Resource
Comments
Medline ID
PubMed ID 11076501
Journal Biochemistry
Year 2000
Volume 39
Pages 13633-40
Authors Gross JW, Hegeman AD, Vestling MM, Frey PA
Title Characterization of enzymatic processes by rapid mix-quench mass spectrometry: the case of dTDP-glucose 4,6-dehydratase.
Related PDB
Related UniProtKB
[8]
Resource
Comments
Medline ID
PubMed ID 11478886
Journal Biochemistry
Year 2001
Volume 40
Pages 9187-95
Authors Gerratana B, Cleland WW, Frey PA
Title Mechanistic roles of Thr134, Tyr160, and Lys 164 in the reaction catalyzed by dTDP-glucose 4,6-dehydratase.
Related PDB
Related UniProtKB
[9]
Resource
Comments
Medline ID
PubMed ID 11601973
Journal Biochemistry
Year 2001
Volume 40
Pages 12497-504
Authors Gross JW, Hegeman AD, Gerratana B, Frey PA
Title Dehydration is catalyzed by glutamate-136 and aspartic acid-135 active site residues in Escherichia coli dTDP-glucose 4,6-dehydratase.
Related PDB
Related UniProtKB
[10]
Resource
Comments
Medline ID
PubMed ID 11380254
Journal Biochemistry
Year 2001
Volume 40
Pages 6598-610
Authors Hegeman AD, Gross JW, Frey PA
Title Probing catalysis by Escherichia coli dTDP-glucose-4,6-dehydratase: identification and preliminary characterization of functional amino acid residues at the active site.
Related PDB
Related UniProtKB
[11]
Resource
Comments
Medline ID
PubMed ID 11553351
Journal Carbohydr Res
Year 2001
Volume 335
Pages 23-32
Authors Amann S, Drager G, Rupprath C, Kirschning A, Elling L
Title (Chemo)enzymatic synthesis of dTDP-activated 2,6-dideoxysugars as building blocks of polyketide antibiotics.
Related PDB
Related UniProtKB
[12]
Resource
Comments X-ray crystallography
Medline ID
PubMed ID 11243820
Journal J Mol Biol
Year 2001
Volume 307
Pages 283-95
Authors Allard ST, Giraud MF, Whitfield C, Graninger M, Messner P, Naismith JH
Title The crystal structure of dTDP-D-Glucose 4,6-dehydratase (RmlB) from Salmonella enterica serovar Typhimurium, the second enzyme in the dTDP-l-rhamnose pathway.
Related PDB 1g1a
Related UniProtKB
[13]
Resource
Comments
Medline ID
PubMed ID 11851427
Journal Biochemistry
Year 2002
Volume 41
Pages 2797-804
Authors Hegeman AD, Gross JW, Frey PA
Title Concerted and stepwise dehydration mechanisms observed in wild-type and mutated Escherichia coli dTDP-glucose 4,6-dehydratase.
Related PDB
Related UniProtKB
[14]
Resource
Comments
Medline ID
PubMed ID 11796113
Journal Structure (Camb)
Year 2002
Volume 10
Pages 81-92
Authors Allard ST, Beis K, Giraud MF, Hegeman AD, Gross JW, Wilmouth RC, Whitfield C, Graninger M, Messner P, Allen AG, Maskell DJ, Naismith JH
Title Toward a structural understanding of the dehydratase mechanism.
Related PDB 1kep 1ker 1keu 1ket 1kew
Related UniProtKB
[15]
Resource
Comments
Medline ID
PubMed ID 14505409
Journal J Am Chem Soc
Year 2003
Volume 125
Pages 11872-8
Authors Beis K, Allard ST, Hegeman AD, Murshudov G, Philp D, Naismith JH
Title The structure of NADH in the enzyme dTDP-d-glucose dehydratase (RmlB).
Related PDB 1oc2
Related UniProtKB
[16]
Resource
Comments
Medline ID
PubMed ID 14570895
Journal J Biol Chem
Year 2004
Volume 279
Pages 2211-20
Authors Allard ST, Cleland WW, Holden HM
Title High resolution X-ray structure of dTDP-glucose 4,6-dehydratase from Streptomyces venezuelae.
Related PDB 1r66 1r6d
Related UniProtKB
[17]
Resource
Comments
Medline ID
PubMed ID 15493979
Journal Biochem Soc Trans
Year 2004
Volume 32
Pages 647-54
Authors Naismith JH
Title Chemical insights from structural studies of enzymes.
Related PDB
Related UniProtKB

Comments
This enzyme belongs to the Short-chain dehydrogenases/reductases (SDR).
Although the catalytic site is slightly different, this enzyme is homologous to the GDP-mannose 4,6-dehydratases (EC 4.2.1.47, D00513 and D00543 in EzCatDB).
This enzyme catalyzes reactions similar to those by its homologous enzymes (D00513 and D00543) (see [14]).
According to the literature [14], [16] and [17], this enzyme catalyzes at least three reactions: oxidation of dTDP-glucose (hydride transfer from dTDP-glucose to nicotinamide), dehydration (elimination of a hydroxyl group from C6), and rereduction of C5-C6 double bond to methyl group (hydride transfer from nicotinamide to the intermediate).
Taken together, this enzyme catalyzes the following reactions:
(A) Hydride transfer from C4 atom of substrate to NAD, forming a 4-keto intermediate (I00091):
(A0) Lys171 (of 1g1a) modulates the activity (or pKa) of Tyr167 via 2'-hydroxyl group of NAD, along with the N1 atom of the nicotinamide group in NAD, whereas Thr133 modulates the pKa of 4-hydroxyl oxygen of the substrate.
(A1) Tyr167 acts as a general base to deprotonate the 4-hydroxyl oxygen of the substrate. Meanwhile, the hydride transfer occurs from the C4-carbon of the substrate to the C4 atom of the nicotinamide, forming the 4-keto intermediate (I00091).
(B) Elimination of hydroxyl group from C6 of the intermediate, forming 4-keto-5,6-glucosen intermediate (I00092):
(B1) Glu135 acts as a general base to deprotonate the C5 atom, possibly forming an enol/enolate transition-state. This transition-state might be stabilized by Tyr167 and Thr133.
(B2) Asp134 acts as a general acid to protonate the O6 hydroxyl group, to release a water molecule, and to form the 4-keto-5,6-ene intermediate from the enol/enolate transition-state.
(C) Hydride transfer from NADH to C6 atom of the intermediate (I00092):
(C0) A slight rotation of the hexose ring of intermediate might be necessary for the reaction. Lys171 modulates the activity (or pKa) of Tyr167 via 2'-hydroxyl group of NAD, along with the N1 atom of the nicotinamide group in NAD.
(C1) Hydride transfer from NADH to C6 atom of the hexose ring in the intermediate. Meanwhile, a general acid must protonate the C5 atom of the hexose. According to the literature [14] and [17], Tyr167 seems to play a role as a general acid. (Thr133 might assist Tyr167 as a general acid, probably acting as a proton shuttle as in the homologous enzyme.)

Created Updated
2004-06-07 2011-06-16