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Complex (oxidoreductase/peptide) PDB id
7r1r
Jmol
Contents
Protein chains
738 a.a. *
18 a.a. *
Ligands
TYR-LEU-VAL-GLY
Waters ×90
* Residue conservation analysis
PDB id:
7r1r
Name: Complex (oxidoreductase/peptide)
Title: Ribonucleotide reductase e441q mutant r1 protein from escherichia coli
Structure: Ribonucleotide reductase r1 protein. Chain: a, b, c. Engineered: yes. Mutation: yes. Ribonucleotide reductase r2 protein. Chain: d, e, f, p. Fragment: c-terminal portion, 20 residues. Engineered: yes
Source: Escherichia coli. Organism_taxid: 562. Gene: nrda. Expressed in: escherichia coli. Expression_system_taxid: 562. Organism_taxid: 562
Biol. unit: Dimer (from PDB file)
Resolution:
3.10Å     R-factor:   0.205     R-free:   0.231
Authors: M.Eriksson,H.Eklund
Key ref:
A.L.Persson et al. (1997). A new mechanism-based radical intermediate in a mutant R1 protein affecting the catalytically essential Glu441 in Escherichia coli ribonucleotide reductase. J Biol Chem, 272, 31533-31541. PubMed id: 9395490 DOI: 10.1074/jbc.272.50.31533
Date:
17-Sep-97     Release date:   18-Mar-98    
PROCHECK
Go to PROCHECK summary
 Headers
 References

Protein chains
Pfam   ArchSchema ?
P00452  (RIR1_ECOLI) -  Ribonucleoside-diphosphate reductase 1 subunit alpha
Seq:
Struc: 		 
Seq:
Struc: 		761 a.a.
738 a.a.*
Protein chains
Pfam   ArchSchema ?
P69924  (RIR2_ECOLI) -  Ribonucleoside-diphosphate reductase 1 subunit beta
Seq:
Struc: 		376 a.a.
18 a.a.
Key:    PfamA domain  Secondary structure  CATH domain
* PDB and UniProt seqs differ at 1 residue position (black cross)

 Enzyme reactions 
   Enzyme class: Chains A, D, B, E, C, F: E.C.1.17.4.1  - Ribonucleoside-diphosphate reductase.
[IntEnz]   [ExPASy]   [KEGG]   [BRENDA]
      Reaction: 2'-deoxyribonucleoside diphosphate + thioredoxin disulfide + H2O = ribonucleoside diphosphate + thioredoxin
2'-deoxyribonucleoside diphosphate
 + thioredoxin disulfide
 + H(2)O
 = ribonucleoside diphosphate
 + thioredoxin
      Cofactor: Iron


Molecule diagrams generated from .mol files obtained from the KEGG ftp site
 Gene Ontology (GO) functional annotation 
  GO annot!
  Cellular component     cytoplasm   2 terms 
  Biological process     oxidation reduction   4 terms 
  Biochemical function     nucleotide binding     5 terms  

 

 
    reference    
 
 
DOI no: 10.1074/jbc.272.50.31533 J Biol Chem 272:31533-31541 (1997)
PubMed id: 9395490  
 
 
A new mechanism-based radical intermediate in a mutant R1 protein affecting the catalytically essential Glu441 in Escherichia coli ribonucleotide reductase.
A.L.Persson, M.Eriksson, B.Katterle, S.Pötsch, M.Sahlin, B.M.Sjöberg.
 
  ABSTRACT  
 
The invariant active site residue Glu441 in protein R1 of ribonucleotide reductase from Escherichia coli has been engineered to alanine, aspartic acid, and glutamic acid. Each mutant protein was structurally and enzymatically characterized. Glu441 contributes to substrate binding, and a carboxylate side chain at position 441 is essential for catalysis. The most intriguing results are the suicidal mechanism-based reaction intermediates observed when R1 E441Q is incubated with protein R2 and natural substrates (CDP and GDP). In a consecutive reaction sequence, we observe at least three clearly discernible steps: (i) a rapid decay (k1 >/= 1.2 s-1) of the catalytically essential tyrosyl radical of protein R2 concomitant with formation of an early transient radical intermediate species, (ii) a slower decay (k2 = 0.03 s-1) of the early intermediate concomitant with formation of another intermediate with a triplet EPR signal, and (iii) decay (k3 = 0.004 s-1) of the latter concomitant with formation of a characteristic substrate degradation product. The characteristics of the triplet EPR signal are compatible with a substrate radical intermediate (most likely localized at the 3'-position of the ribose moiety of the substrate nucleotide) postulated to occur in the wild type reaction mechanism as well.
 
  Selected figure(s)  
 
Figure 1.
Fig. 1. A, structure of substrate binding at the active site of reduced protein R1. The 3 -oxygen is hydrogen-bonded to the side chain of Glu441, the 2 -oxygen is hydrogen-bonded to the side chain of Asn437 and Cys225, and Glu441 and Asn437 are hydrogen-bonded. These interactions are indicated by thin lines. The postulated hydrogen-bonded radical transfer pathway is indicated by dashed lines. Adapted from Eriksson et al. (16). B, stereopairs of the structure of the active site environment of the three mutant R1 proteins and wild type R1. The mutant residues at position 441, Asn437, Cys225, Cys439, Cys462 and Met620 are indicated. Wild type, green; E441A, blue; E441D, red; E441Q, yellow. 		Figure 2.
Fig. 2. Proposed reaction mechanism for reduction of ribonucleotides by ribonucleotide reductase. In 3, two alternative^ reduction pathways have been indicated, a and b1-b2, respectively.
 
  The above figures are reprinted by permission from the ASBMB: J Biol Chem (1997, 272, 31533-31541) copyright 1997.  
  Figures were selected by an automated process.  

Literature references that cite this PDB file's key reference

  PubMed id Reference
19128178 H.Zipse, E.Artin, S.Wnuk, G.J.Lohman, D.Martino, R.G.Griffin, S.Kacprzak, M.Kaupp, B.Hoffman, M.Bennati, J.Stubbe, and N.Lees (2009).
Structure of the nucleotide radical formed during reaction of CDP/TTP with the E441Q-alpha2beta2 of E. coli ribonucleotide reductase.
  J Am Chem Soc, 131, 200-211.  
16361339 N.M.Cerqueira, P.A.Fernandes, L.A.Eriksson, and M.J.Ramos (2006).
Dehydration of ribonucleotides catalyzed by ribonucleotide reductase: the role of the enzyme.
  Biophys J, 90, 2109-2119.  
16261381 S.Pereira, N.M.Cerqueira, P.A.Fernandes, and M.J.Ramos (2006).
Computational studies on class I ribonucleotide reductase: understanding the mechanisms of action and inhibition of a cornerstone enzyme for the treatment of cancer.
  Eur Biophys J, 35, 125-135.  
16218873 M.Bennati, F.Lendzian, M.Schmittel, and H.Zipse (2005).
Spectroscopic and theoretical approaches for studying radical reactions in class I ribonucleotide reductase.
  Biol Chem, 386, 1007-1022.  
15481089 N.M.Cerqueira, P.A.Fernandes, L.A.Eriksson, and M.J.Ramos (2004).
Ribonucleotide activation by enzyme ribonucleotide reductase: understanding the role of the enzyme.
  J Comput Chem, 25, 2031-2037.  
14648621 S.Pereira, P.A.Fernandes, and M.J.Ramos (2004).
Theoretical study of ribonucleotide reductase mechanism-based inhibition by 2'-azido-2'-deoxyribonucleoside 5'-diphosphates.
  J Comput Chem, 25, 227-237.  
15139041 S.Pereira, P.A.Fernandes, and M.J.Ramos (2004).
Mechanism for ribonucleotide reductase inactivation by the anticancer drug gemcitabine.
  J Comput Chem, 25, 1286-1294.  
14696066 V.Pelmenschikov, K.B.Cho, and P.E.Siegbahn (2004).
Class I ribonucleotide reductase revisited: the effect of removing a proton on Glu441.
  J Comput Chem, 25, 311-321.  
12562785 M.Ekberg, P.Birgander, and B.M.Sjöberg (2003).
In vivo assay for low-activity mutant forms of Escherichia coli ribonucleotide reductase.
  J Bacteriol, 185, 1167-1173.  
10769119 O.Guittet, P.Decottignies, L.Serani, Y.Henry, P.Le Maréchal, O.Laprévote, and M.Lepoivre (2000).
Peroxynitrite-mediated nitration of the stable free radical tyrosine residue of the ribonucleotide reductase small subunit.
  Biochemistry, 39, 4640-4648.  
10430881 C.C.Lawrence, M.Bennati, H.V.Obias, G.Bar, R.G.Griffin, and J.Stubbe (1999).
High-field EPR detection of a disulfide radical anion in the reduction of cytidine 5'-diphosphate by the E441Q R1 mutant of Escherichia coli ribonucleotide reductase.
  Proc Natl Acad Sci U S A, 96, 8979-8984.  
  10574800 H.Eklund, and M.Fontecave (1999).
Glycyl radical enzymes: a conservative structural basis for radicals.
  Structure, 7, R257-R262.  
10542051 S.Sauge-Merle, D.Falconet, and M.Fontecave (1999).
An active ribonucleotide reductase from Arabidopsis thaliana cloning, expression and characterization of the large subunit.
  Eur J Biochem, 266, 62-69.  
9501154 J.Stubbe (1998).
Ribonucleotide reductases in the twenty-first century.
  Proc Natl Acad Sci U S A, 95, 2723-2724.  
  9852763 J.Stubbe, and P.Riggs-Gelasco (1998).
Harnessing free radicals: formation and function of the tyrosyl radical in ribonucleotide reductase.
  Trends Biochem Sci, 23, 438-443.  
9572859 W.A.van der Donk, G.Yu, L.Pérez, R.J.Sanchez, J.Stubbe, V.Samano, and M.J.Robins (1998).
Detection of a new substrate-derived radical during inactivation of ribonucleotide reductase from Escherichia coli by gemcitabine 5'-diphosphate.
  Biochemistry, 37, 6419-6426.  
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.