DB code: S00067

CATH domain	2.30.30.60 : SH3 type barrels.	Catalytic domain
E.C.	1.5.1.3
CSA	1vie
M-CSA	1vie
MACiE

CATH domain	Related DB codes (homologues)

Uniprot Enzyme Name
UniprotKB	Protein name	Synonyms	Pfam
P00383	Dihydrofolate reductase type 2	EC 1.5.1.3 Dihydrofolate reductase type II	PF06442 (DHFR_2) [Graphical View]

KEGG enzyme name
dihydrofolate reductase tetrahydrofolate dehydrogenase DHFR pteridine reductase:dihydrofolate reductase dihydrofolate reductase:thymidylate synthase thymidylate synthetase-dihydrofolate reductase folic acid reductase folic reductase dihydrofolic acid reductase dihydrofolic reductase 7,8-dihydrofolate reductase NADPH-dihydrofolate reductase

UniprotKB: Accession Number	Entry name	Activity	Subunit	Subcellular location	Cofactor
P00383	DYR21_ECOLX	5,6,7,8-tetrahydrofolate + NADP(+) = 7,8- dihydrofolate + NADPH.

KEGG Pathways
Map code	Pathways	E.C.
MAP00670	One carbon pool by folate
MAP00790	Folate biosynthesis

Compound table
	Substrates			Products		Intermediates
KEGG-id	C00415	C00005	C00080	C00101	C00006
E.C.
Compound	7,8-Dihydrofolate	NADPH	H+	5,6,7,8-Tetrahydrofolate	NADP+
Type	amino acids,amide group,amine group,aromatic ring (only carbon atom),aromatic ring (with nitrogen atoms),carboxyl group	amide group,amine group,nucleotide	others	amino acids,amide group,amine group,aromatic ring (only carbon atom),aromatic ring (with nitrogen atoms),carboxyl group	amide group,amine group,nucleotide
ChEBI	15633 15633	16474 16474	15378 15378	15635 20506 15635 20506	18009 18009
PubChem	98792 98792	5884 5884	1038 1038	5460413 91443 5460413 91443	5886 5886

1vieA	Unbound	Unbound		Unbound	Unbound
1vifA	Analogue:FOL	Unbound		Unbound	Unbound
2gqvA	Unbound	Unbound		Unbound	Unbound
2p4tA	Unbound	Unbound		Unbound	Analogue:NAP
2rh2A	Unbound	Unbound		Unbound	Unbound
2rk1A	Analogue:DHF	Unbound		Unbound	Bound:NAP
2rk2A	Unbound	Unbound		Unbound	Bound:NAP

Reference for Active-site residues
resource	references	E.C.

Active-site residues
PDB	Catalytic residues	Cofactor-binding residues	Modified residues	Main-chain involved in catalysis	Comment

1vieA				ILE 68
1vifA				ILE 68
2gqvA				ILE 68
2p4tA				ILE 68	mutant Q67H
2rh2A				ILE 68
2rk1A				ILE 68
2rk2A				ILE 68

References for Catalytic Mechanism
References	Sections	No. of steps in catalysis
[1]	p.4201-4202
[4]
[11]
[12]	p.1048-1049
[13]
[16]	p.14485-14486, Fig.7

References
[1]
Resource
Comments
Medline ID
PubMed ID	3530319
Journal	Biochemistry
Year	1986
Volume	25
Pages	4194-204
Authors	Matthews DA, Smith SL, Baccanari DP, Burchall JJ, Oatley SJ, Kraut J
Title	Crystal structure of a novel trimethoprim-resistant dihydrofolate reductase specified in Escherichia coli by R-plasmid R67.
Related PDB
Related UniProtKB
[2]
Resource
Comments
Medline ID
PubMed ID	1932013
Journal	Biochemistry
Year	1991
Volume	30
Pages	10895-904
Authors	Reece LJ, Nichols R, Ogden RC, Howell EE
Title	Construction of a synthetic gene for an R-plasmid-encoded dihydrofolate reductase and studies on the role of the N-terminus in the protein.
Related PDB
Related UniProtKB
[3]
Resource
Comments
Medline ID
PubMed ID	8226776
Journal	J Biol Chem
Year	1993
Volume	268
Pages	22672-9
Authors	Zhuang P, Yin M, Holland JC, Peterson CB, Howell EE
Title	Artificial duplication of the R67 dihydrofolate reductase gene to create protein asymmetry. Effects on protein activity and folding.
Related PDB
Related UniProtKB
[4]
Resource
Comments	X-RAY CRYSTALLOGRAPHY (1.7 ANGSTROMS) OF 17-78
Medline ID	96069790
PubMed ID	7583655
Journal	Nat Struct Biol
Year	1995
Volume	2
Pages	1018-25
Authors	Narayana N, Matthews DA, Howell EE, Nguyen-huu X
Title	A plasmid-encoded dihydrofolate reductase from trimethoprim-resistant bacteria has a novel D2-symmetric active site.
Related PDB	1vie 1vif
Related UniProtKB	P00383
[5]
Resource
Comments
Medline ID
PubMed ID	8784197
Journal	Biochemistry
Year	1996
Volume	35
Pages	11414-24
Authors	Bradrick TD, Beechem JM, Howell EE
Title	Unusual binding stoichiometries and cooperativity are observed during binary and ternary complex formation in the single active pore of R67 dihydrofolate reductase, a D2 symmetric protein.
Related PDB
Related UniProtKB
[6]
Resource
Comments
Medline ID
PubMed ID	8910413
Journal	J Biol Chem
Year	1996
Volume	271
Pages	28031-7
Authors	Bradrick TD, Shattuck C, Strader MB, Wicker C, Eisenstein E, Howell EE
Title	Redesigning the quaternary structure of R67 dihydrofolate reductase. Creation of an active monomer from a tetrameric protein by quadruplication of the gene.
Related PDB
Related UniProtKB
[7]
Resource
Comments
Medline ID
PubMed ID	8999931
Journal	J Biol Chem
Year	1997
Volume	272
Pages	2252-8
Authors	Park H, Zhuang P, Nichols R, Howell EE
Title	Mechanistic studies of R67 dihydrofolate reductase. Effects of pH and an H62C mutation.
Related PDB
Related UniProtKB
[8]
Resource
Comments
Medline ID
PubMed ID	9543003
Journal	Protein Eng
Year	1997
Volume	10
Pages	1415-24
Authors	Park H, Bradrick TD, Howell EE
Title	A glutamine 67--> histidine mutation in homotetrameric R67 dihydrofolate reductase results in four mutations per single active site pore and causes substantial substrate and cofactor inhibition.
Related PDB
Related UniProtKB
[9]
Resource
Comments
Medline ID
PubMed ID	10964572
Journal	J Mol Biol
Year	2000
Volume	302
Pages	235-50
Authors	Dam J, Rose T, Goldberg ME, Blondel A
Title	Complementation between dimeric mutants as a probe of dimer-dimer interactions in tetrameric dihydrofolate reductase encoded by R67 plasmid of E. coli.
Related PDB
Related UniProtKB
[10]
Resource
Comments
Medline ID
PubMed ID	11284680
Journal	Biochemistry
Year	2001
Volume	40
Pages	4242-52
Authors	Li D, Levy LA, Gabel SA, Lebetkin MS, DeRose EF, Wall MJ, Howell EE, London RE
Title	Interligand Overhauser effects in type II dihydrofolate reductase.
Related PDB
Related UniProtKB
[11]
Resource
Comments
Medline ID
PubMed ID	11560482
Journal	Biochemistry
Year	2001
Volume	40
Pages	11344-52
Authors	Strader MB, Smiley RD, Stinnett LG, VerBerkmoes NC, Howell EE
Title	Role of S65, Q67, I68, and Y69 residues in homotetrameric R67 dihydrofolate reductase.
Related PDB
Related UniProtKB
[12]
Resource
Comments
Medline ID
PubMed ID	11989624
Journal	J Comput Aided Mol Des
Year	2001
Volume	15
Pages	1035-52
Authors	Howell EE, Shukla U, Hicks SN, Smiley RD, Kuhn LA, Zavodszky MI
Title	One site fits both: a model for the ternary complex of folate + NADPH in R67 dihydrofolate reductase, a D2 symmetric enzyme.
Related PDB
Related UniProtKB
[13]
Resource
Comments
Medline ID
PubMed ID	15812782
Journal	Chembiochem
Year	2005
Volume	6
Pages	590-600
Authors	Howell EE
Title	Searching sequence space: two different approaches to dihydrofolate reductase catalysis.
Related PDB
Related UniProtKB
[14]
Resource
Comments
Medline ID
PubMed ID	16790925
Journal	Acta Crystallogr D Biol Crystallogr
Year	2006
Volume	62
Pages	695-706
Authors	Narayana N
Title	High-resolution structure of a plasmid-encoded dihydrofolate reductase: pentagonal network of water molecules in the D2-symmetric active site.
Related PDB	2gqv
Related UniProtKB
[15]
Resource
Comments
Medline ID
PubMed ID	17473013
Journal	Protein Sci
Year	2007
Volume	16
Pages	1063-8
Authors	Divya N, Grifith E, Narayana N
Title	Structure of the Q67H mutant of R67 dihydrofolate reductase-NADP+ complex reveals a novel cofactor binding mode.
Related PDB	2p4t
Related UniProtKB
[16]
Resource
Comments
Medline ID
PubMed ID	18052202
Journal	Biochemistry
Year	2007
Volume	46
Pages	14878-88
Authors	Krahn JM, Jackson MR, DeRose EF, Howell EE, London RE
Title	Crystal structure of a type II dihydrofolate reductase catalytic ternary complex.
Related PDB	2rh2 2rk1 2rk2
Related UniProtKB

Comments
According to the literature [13], Lys32, Gln67, and Tyr69 have been reported to act as catalytic residues. However, they seem to contribute to the ligand binding, rather than catalysis (see [16]). According to the literature [16], the reaction of this enzyme occurs in two steps: a hydride transfer from NADPH to the C6 atom of dihydrofolate (DHF), and a protonation step. The protonation step is more likely to preceed the hydride transfer step (see [16]), in general. However, no residues in close to the ligand seem to be involved in the protonation. On the other hand, the p-aminobenzoyl glutamate tail of DHF might be involved in substrate-assisted catalysis (see [16]). Its glutamate group might donate the proton to the N5 atom of DHF, to facilitate the reaction. In the next step, the hydride transfer may be assisted by the mainchain amide and carbonyl of Ile68 (see [16]).

Comments

According to the literature [13], Lys32, Gln67, and Tyr69 have been reported to act as catalytic residues. However, they seem to contribute to the ligand binding, rather than catalysis (see [16]).
According to the literature [16], the reaction of this enzyme occurs in two steps: a hydride transfer from NADPH to the C6 atom of dihydrofolate (DHF), and a protonation step. The protonation step is more likely to preceed the hydride transfer step (see [16]), in general. However, no residues in close to the ligand seem to be involved in the protonation. On the other hand, the p-aminobenzoyl glutamate tail of DHF might be involved in substrate-assisted catalysis (see [16]). Its glutamate group might donate the proton to the N5 atom of DHF, to facilitate the reaction. In the next step, the hydride transfer may be assisted by the mainchain amide and carbonyl of Ile68 (see [16]).

Created	Updated
2004-01-29	2009-03-17