PDBsum entry 1j79

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protein ligands metals Protein-protein interface(s) links
Hydrolase PDB id
Protein chains
343 a.a. *
NCD ×2
_ZN ×4
Waters ×731
* Residue conservation analysis
PDB id:
Name: Hydrolase
Title: Molecular structure of dihydroorotase: a paradigm for catalysis through the use of a binuclear metal center
Structure: Dihydroorotase. Chain: a, b. Synonym: dhoase. Engineered: yes
Source: Escherichia coli. Organism_taxid: 562. Gene: pyrc. Expressed in: escherichia coli. Expression_system_taxid: 562
Biol. unit: Dimer (from PQS)
1.70Å     R-factor:   0.193     R-free:   0.258
Authors: J.B.Thoden,G.N.Phillips Jr.,T.M.Neal,F.M.Raushel,H.M.Holden
Key ref:
J.B.Thoden et al. (2001). Molecular structure of dihydroorotase: a paradigm for catalysis through the use of a binuclear metal center. Biochemistry, 40, 6989-6997. PubMed id: 11401542
16-May-01     Release date:   20-Jun-01    
Go to PROCHECK summary

Protein chains
Pfam   ArchSchema ?
P05020  (PYRC_ECOLI) -  Dihydroorotase
348 a.a.
343 a.a.*
Key:    PfamA domain  Secondary structure  CATH domain
* PDB and UniProt seqs differ at 4 residue positions (black crosses)

 Enzyme reactions 
   Enzyme class: E.C.  - Dihydroorotase.
[IntEnz]   [ExPASy]   [KEGG]   [BRENDA]

Pyrimidine Biosynthesis
      Reaction: (S)-dihydroorotate + H2O = N-carbamoyl-L-aspartate
Bound ligand (Het Group name = ORO)
corresponds exactly
+ H(2)O
Bound ligand (Het Group name = NCD)
corresponds exactly
Molecule diagrams generated from .mol files obtained from the KEGG ftp site
 Gene Ontology (GO) functional annotation 
  GO annot!
  Biological process     'de novo' UMP biosynthetic process   4 terms 
  Biochemical function     hydrolase activity     5 terms  


    Added reference    
Biochemistry 40:6989-6997 (2001)
PubMed id: 11401542  
Molecular structure of dihydroorotase: a paradigm for catalysis through the use of a binuclear metal center.
J.B.Thoden, G.N.Phillips, T.M.Neal, F.M.Raushel, H.M.Holden.
Dihydroorotase plays a key role in pyrimidine biosynthesis by catalyzing the reversible interconversion of carbamoyl aspartate to dihydroorotate. Here we describe the three-dimensional structure of dihydroorotase from Escherichia coli determined and refined to 1.7 A resolution. Each subunit of the homodimeric enzyme folds into a "TIM" barrel motif with eight strands of parallel beta-sheet flanked on the outer surface by alpha-helices. Unexpectedly, each subunit contains a binuclear zinc center with the metal ions separated by approximately 3.6 A. Lys 102, which is carboxylated, serves as a bridging ligand between the two cations. The more buried or alpha-metal ion in subunit I is surrounded by His 16, His 18, Lys 102, Asp 250, and a solvent molecule (most likely a hydroxide ion) in a trigonal bipyramidal arrangement. The beta-metal ion, which is closer to the solvent, is tetrahedrally ligated by Lys 102, His 139, His 177, and the bridging hydroxide. L-Dihydroorotate is observed bound to subunit I, with its carbonyl oxygen, O4, lying 2.9 A from the beta-metal ion. Important interactions for positioning dihydroorotate into the active site include a salt bridge with the guanidinium group of Arg 20 and various additional electrostatic interactions with both protein backbone and side chain atoms. Strikingly, in subunit II, carbamoyl L-aspartate is observed binding near the binuclear metal center with its carboxylate side chain ligating the two metals and thus displacing the bridging hydroxide ion. From the three-dimensional structures of the enzyme-bound substrate and product, it has been possible to propose a unique catalytic mechanism for dihydroorotase. In the direction of dihydroorotate hydrolysis, the bridging hydroxide attacks the re-face of dihydroorotate with general base assistance by Asp 250. The carbonyl group is polarized for nucleophilic attack by the bridging hydroxide through a direct interaction with the beta-metal ion. During the cyclization of carbamoyl aspartate, Asp 250 initiates the reaction by abstracting a proton from N3 of the substrate. The side chain carboxylate of carbamoyl aspartate is polarized through a direct electrostatic interaction with the binuclear metal center. The ensuing tetrahedral intermediate collapses with C-O bond cleavage and expulsion of the hydroxide which then bridges the binuclear metal center.

Literature references that cite this PDB file's key reference

  PubMed id Reference
20676924 C.C.Wang, H.W.Tsau, W.T.Chen, and C.Y.Huang (2010).
Identification and characterization of a putative dihydroorotase, KPN01074, from Klebsiella pneumoniae.
  Protein J, 29, 445-452.  
21143781 C.L.Hung, C.Lee, C.Y.Lin, C.H.Chang, Y.C.Chung, and C.Yi Tang (2010).
Feature amplified voting algorithm for functional analysis of protein superfamily.
  BMC Genomics, 11, S14.  
20000809 J.A.Cummings, T.T.Nguyen, A.A.Fedorov, P.Kolb, C.Xu, E.V.Fedorov, B.K.Shoichet, D.P.Barondeau, S.C.Almo, and F.M.Raushel (2010).
Structure, mechanism, and substrate profile for Sco3058: the closest bacterial homologue to human renal dipeptidase .
  Biochemistry, 49, 611-622.
PDB codes: 3id7 3itc 3k5x
20386924 V.Kõiv, L.Andresen, and A.Mäe (2010).
AepA of Pectobacterium is not involved in the regulation of extracellular plant cell wall degrading enzymes production.
  Mol Genet Genomics, 283, 541-549.  
19822009 C.E.Haas, D.A.Rodionov, J.Kropat, D.Malasarn, S.S.Merchant, and Crécy-Lagard (2009).
A subset of the diverse COG0523 family of putative metal chaperones is linked to zinc homeostasis in all kingdoms of life.
  BMC Genomics, 10, 470.  
18781344 C.Y.Huang, C.C.Hsu, M.C.Chen, and Y.S.Yang (2009).
Effect of metal binding and posttranslational lysine carboxylation on the activity of recombinant hydantoinase.
  J Biol Inorg Chem, 14, 111-121.  
19241382 E.M.Meulenbroek, K.Paspaleva, E.A.Thomassen, J.P.Abrahams, N.Goosen, and N.S.Pannu (2009).
Involvement of a carboxylated lysine in UV damage endonuclease.
  Protein Sci, 18, 549-558.
PDB codes: 3bzg 3bzj 3c0l 3c0q 3c0s
18702530 J.Kim, P.C.Tsai, S.L.Chen, F.Himo, S.C.Almo, and F.M.Raushel (2008).
Structure of diethyl phosphate bound to the binuclear metal center of phosphotriesterase.
  Biochemistry, 47, 9497-9504.
PDB codes: 2o4q 3cak 3cs2
18282099 S.Samant, H.Lee, M.Ghassemi, J.Chen, J.L.Cook, A.S.Mankin, and A.A.Neyfakh (2008).
Nucleotide biosynthesis is critical for growth of bacteria in human blood.
  PLoS Pathog, 4, e37.  
17124633 L.Niu, X.Zhang, Y.Shi, and J.Yuan (2007).
Subunit dissociation and stability alteration of D hydantoinase deleted at the terminal amino acid residue.
  Biotechnol Lett, 29, 303-308.  
  17329804 M.Lee, M.J.Maher, and J.M.Guss (2007).
Structure of the T109S mutant of Escherichia coli dihydroorotase complexed with the inhibitor 5-fluoroorotate: catalytic activity is reflected by the crystal form.
  Acta Crystallogr Sect F Struct Biol Cryst Commun, 63, 154-161.
PDB code: 2e25
17567047 R.S.Hall, D.F.Xiang, C.Xu, and F.M.Raushel (2007).
N-Acetyl-D-glucosamine-6-phosphate deacetylase: substrate activation via a single divalent metal ion.
  Biochemistry, 46, 7942-7952.  
16517602 B.Lohkamp, B.Andersen, J.Piskur, and D.Dobritzsch (2006).
The crystal structures of dihydropyrimidinases reaffirm the close relationship between cyclic amidohydrolases and explain their substrate specificity.
  J Biol Chem, 281, 13762-13776.
PDB codes: 2ftw 2fty 2fvk 2fvm
16963440 M.Goto, H.Hayashi, I.Miyahara, K.Hirotsu, M.Yoshida, and T.Oikawa (2006).
Crystal structures of nonoxidative zinc-dependent 2,6-dihydroxybenzoate (gamma-resorcylate) decarboxylase from Rhizobium sp. strain MTP-10005.
  J Biol Chem, 281, 34365-34373.
PDB codes: 2dvt 2dvu 2dvx
15521054 F.Bramsen, A.D.Bond, C.J.McKenzie, R.G.Hazell, B.Moubaraki, and K.S.Murray (2005).
Self-assembly of the octanuclear cluster [Cu8(OH)10(NH2(CH2)2CH3)12]6+ and the one-dimensional N-propylcarbamate-linked coordination polymer {[Cu(O2CNH(CH2)2CH3)(NH2(CH2)2CH3)3](ClO4)}n.
  Chemistry, 11, 825-831.  
16199586 J.K.Kim, S.B.Mulrooney, and R.P.Hausinger (2005).
Biosynthesis of active Bacillus subtilis urease in the absence of known urease accessory proteins.
  J Bacteriol, 187, 7150-7154.  
16121396 J.Li, J.B.Cross, T.Vreven, S.O.Meroueh, S.Mobashery, and H.B.Schlegel (2005).
Lysine carboxylation in proteins: OXA-10 beta-lactamase.
  Proteins, 61, 246-257.  
16284925 L.L.Lin, W.H.Hsu, W.Y.Hsu, S.C.Kan, and H.Y.Hu (2005).
Phylogenetic analysis and biochemical characterization of a thermostable dihydropyrimidinase from alkaliphilic Bacillus sp. TS-23.
  Antonie Van Leeuwenhoek, 88, 189-197.  
16211624 S.H.Nam, H.S.Park, and H.S.Kim (2005).
Evolutionary relationship and application of a superfamily of cyclic amidohydrolase enzymes.
  Chem Rec, 5, 298-307.  
15381710 A.Ahuja, C.Purcarea, R.Ebert, S.Sadecki, H.I.Guy, and D.R.Evans (2004).
Aquifex aeolicus dihydroorotase: association with aspartate transcarbamoylase switches on catalytic activity.
  J Biol Chem, 279, 53136-53144.  
15278241 D.M.Brichta, K.N.Azad, P.Ralli, and G.A.O'Donovan (2004).
Pseudomonas aeruginosa dihydroorotases: a tale of three pyrCs.
  Arch Microbiol, 182, 7.  
15096496 D.R.Evans, and H.I.Guy (2004).
Mammalian pyrimidine biosynthesis: fresh insights into an ancient pathway.
  J Biol Chem, 279, 33035-33038.  
15358859 H.Fukatsu, Y.Hashimoto, M.Goda, H.Higashibata, and M.Kobayashi (2004).
Amine-synthesizing enzyme N-substituted formamide deformylase: screening, purification, characterization, and gene cloning.
  Proc Natl Acad Sci U S A, 101, 13726-13731.  
14685275 R.C.Deo, E.F.Schmidt, A.Elhabazi, H.Togashi, S.K.Burley, and S.M.Strittmatter (2004).
Structural bases for CRMP function in plexin-dependent semaphorin3A signaling.
  EMBO J, 23, 9.
PDB code: 1kcx
14736882 W.L.Lai, L.Y.Chou, C.Y.Ting, R.Kirby, Y.C.Tsai, A.H.Wang, and S.H.Liaw (2004).
The functional role of the binuclear metal center in D-aminoacylase: one-metal activation and second-metal attenuation.
  J Biol Chem, 279, 13962-13967.
PDB codes: 1rjp 1rjq 1rjr 1rk5 1rk6 1v4y 1v51
12554960 M.J.Maher, D.T.Huang, J.M.Guss, C.A.Collyer, and R.I.Christopherson (2003).
Crystallization of hamster dihydroorotase: involvement of a disulfide-linked tetrameric form.
  Acta Crystallogr D Biol Crystallogr, 59, 381-384.  
12486048 S.B.Mulrooney, and R.P.Hausinger (2003).
Metal ion dependence of recombinant Escherichia coli allantoinase.
  J Bacteriol, 185, 126-134.  
12454005 S.H.Liaw, S.J.Chen, T.P.Ko, C.S.Hsu, C.J.Chen, A.H.Wang, and Y.C.Tsai (2003).
Crystal structure of D-aminoacylase from Alcaligenes faecalis DA1. A novel subset of amidohydrolases and insights into the enzyme mechanism.
  J Biol Chem, 278, 4957-4962.
PDB code: 1m7j
12441349 S.Mangani, W.Meyer-Klaucke, A.J.Moir, M.Ranieri-Raggi, D.Martini, and A.Raggi (2003).
Characterization of the zinc-binding site of the histidine-proline-rich glycoprotein associated with rabbit skeletal muscle AMP deaminase.
  J Biol Chem, 278, 3176-3184.  
12626710 Z.Gojkovic, L.Rislund, B.Andersen, M.P.Sandrini, P.F.Cook, K.D.Schnackerz, and J.Piskur (2003).
Dihydropyrimidine amidohydrolases and dihydroorotases share the same origin and several enzymatic properties.
  Nucleic Acids Res, 31, 1683-1692.  
12837777 Z.Xu, Y.Liu, Y.Yang, W.Jiang, E.Arnold, and J.Ding (2003).
Crystal structure of D-Hydantoinase from Burkholderia pickettii at a resolution of 2.7 Angstroms: insights into the molecular basis of enzyme thermostability.
  J Bacteriol, 185, 4038-4049.
PDB code: 1nfg
12136154 B.Beuth, K.Niefind, and D.Schomburg (2002).
Crystallization and preliminary crystallographic analysis of creatininase from Pseudomonas putida.
  Acta Crystallogr D Biol Crystallogr, 58, 1356-1358.  
11748240 C.L.Soong, J.Ogawa, E.Sakuradani, and S.Shimizu (2002).
Barbiturase, a novel zinc-containing amidohydrolase involved in oxidative pyrimidine metabolism.
  J Biol Chem, 277, 7051-7058.  
12381838 C.S.Hsu, W.L.Lai, W.W.Chang, S.H.Liaw, and Y.C.Tsai (2002).
Structural-based mutational analysis of D-aminoacylase from Alcaligenes faecalis DA1.
  Protein Sci, 11, 2545-2550.  
12057683 F.M.Raushel (2002).
Bacterial detoxification of organophosphate nerve agents.
  Curr Opin Microbiol, 5, 288-295.  
12056889 M.Sundd, N.Iverson, B.Ibarra-Molero, J.M.Sanchez-Ruiz, and A.D.Robertson (2002).
Electrostatic interactions in ubiquitin: stabilization of carboxylates by lysine amino groups.
  Biochemistry, 41, 7586-7596.  
11849938 J.Altenbuchner, M.Siemann-Herzberg, and C.Syldatk (2001).
Hydantoinases and related enzymes as biocatalysts for the synthesis of unnatural chiral amino acids.
  Curr Opin Biotechnol, 12, 559-563.  
The most recent references are shown first. Citation data come partly from CiteXplore and partly from an automated harvesting procedure. Note that this is likely to be only a partial list as not all journals are covered by either method. However, we are continually building up the citation data so more and more references will be included with time. Where a reference describes a PDB structure, the PDB codes are shown on the right.