Ribonucleotide Reduction

Abstract

Ribonucleotide reductase is an essential enzyme that supplies the deoxyribonucleotides required for DNA synthesis and repair. This one enzyme reduces all four ribonucleotides to the corresponding deoxynucleotides. The enzyme uses a complex radical‐based mechanism to catalyse this reaction. Three main classes of enzymes have been described that differ mainly in the type of cofactor used to generate the catalytic radical. Despite their common reaction mechanism, ribonucleotide reductases show little sequence identity. Crystal structures of enzymes from all classes show a conserved β/α barrel structure in the catalytic domain. Reductase activity is regulated at the protein level by nucleotide allosteric effectors to produce balanced pools of deoxynucleotides. Reductase activity is also coordinated with the cell cycle by activation and repression of gene expression.

Key Concepts

  • Deoxyribonucleotides required for DNA synthesis and repair are generated only by reduction of ribonucleotides.
  • One enzyme, ribonucleotide reductase, must reduce all four common ribonucleotides and generate a balanced pool of deoxyribonucleotides to avoid mutations.
  • Although an essential enzyme in all divisions of life, ribonucleotide reductases diverge widely in primary structure and cofactor requirements.
  • Ribonucleotide reductase is regulated both at the protein and gene level and can accommodate to environmental changes in the evolution of diverse organisms.
  • Evolution of ribonucleotide reductases is constrained only by basic catalytic chemistry and protein tertiary structure.

Keywords: adenosylcobalamin; alpha/beta barrel; ATP cone; deoxynucleotide pools; glycyl radical; thioredoxin; thiyl radical; tyrosyl radical

Figure 3. Structure of the β2 (R2) protein of class Ia ribonucleotide reductase from . The β‐subunits are shown in pink and orange. The structure is mainly helices with a small section of four β‐strands at the tip. The di‐iron centres in each subunit are red spheres bridged by a blue oxygen and are buried within the helical bundles. No radical was present in this structure. The tyrosine that will form a stable radical is shown in yellow and is located near the di‐iron centre. The figure was generated from pdb file 1RIB with Pymol.
Figure 4. Structure of one of the α‐subunits in class Ia ribonucleotide reductase. The 10‐stranded β/α barrel domain is in the centre of the figure. Loop 2 is in the centre of the barrel with the catalytic‐active cysteine shown as a yellow stick figure. The redox‐active cysteines are shown in orange and are present as a disulphide‐bonded cystine in this structure. The glutamic acid that acts as an acid/base in catalysis is shown in red. An ATP analogue is shown as a space‐filling structure and is bound to the ATP cone domain near the ‐terminal of the protein. The figure was generated from pdb file 3R1R with Pymol.
Figure 5. Structure of the dimeric class II ribonucleotide reductase from . The subunit on the right has a bound adenosylcobalamin in a space‐filling structure. The subunit on the left shows the cofactor as a stick figure in red with the central cobalt atom in blue. The catalytic cysteine residue is magenta and is located on a loop in the β/α barrel. The substrate, GDP, is a yellow stick figure also in the active site. The specificity effector, dTTP is bound at the subunit interface and is shown in blue. The figure was generated from pdb file 1XJE with Pymol.
Figure 6. Structure of the dATP‐inhibited tetrameric form of the class Ia ribonucleotide reductase from . The dimeric α‐proteins are shown in blue and green. The dimeric β‐proteins are in red and orange. Orange space‐filling structures are dATP molecules that are bound to the ATP cones of the α‐subunits. The structure shows the unnatural binding of the β2 subunits, which normally fit into the groove between the α‐subunits at the sites shown by orange and red peptides in each α‐structure. The structure was generated from pdb file 3UUS with Pymol.
Figure 1. Reduction of a ribonucleotide to a 2′‐deoxyribonucleotide catalysed by ribonucleotide reductase. R = mono‐ or di‐phosphoester.
Figure 2. Mechanism of ribonucleotide reduction showing the activity of three cysteine residues and a glutamic acid in the active site. B – purine or pyrimidine base. PP – di‐ or tri‐phosphoester. (Modified from Nordlund and Reichard, 2006 © Annual Reviews.)
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Further Reading

Brignole EJ, Ando N, Zimanyi CM and Drennan CL (2012) The prototypic class Ia ribonucleotide reductase from Escherichia coli: still surprising after all these years. Biochemical Society Transactions 40: 523–530.

Cotruvo JA and Stubbe J (2011) Class I ribonucleotide reductases: metallocofactor assembly and repair in vitro and in vivo. Annual Review of Biochemistry 80: 733–767.

Dwivedi B, Xue B, Lundin D, et al. (2013) A bioinformatics analysis of ribonucleotide reductase genes in phage genomes and metagenomes. BMC Evolutionary Biology 13: 33.

Eklund H, Uhlin U, Färnegårdh M, et al. (2001) Structure and function of the radical enzyme ribonucleotide reductase. Progress in Biophysics and Molecular Biology 77: 177–268.

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Huang M, Parker MJ and Stubbe J (2014) Choosing the right metal: case studies of class I ribonucleotide reductases. Journal of Biological Chemistry 289 (41): 28104–28111.

Lundin D, Gribaldo S, Torrents E, et al. (2010) Ribonucleotide reduction – horizontal transfer of a required function spans all three domains. BMC Evolutionary Biology 10: 383.

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Gleason, Florence K(Mar 2015) Ribonucleotide Reduction. In: eLS. John Wiley & Sons Ltd, Chichester. http://www.els.net [doi: 10.1002/9780470015902.a0000567.pub3]