Radical S‐Adenosylmethionine (SAM) Superfamily


A large superfamily of enzymes uses S‐adenosyl‐l‐methionine (SAM) to generate high‐energy carbon radicals as intermediates in a variety of metabolic and biosynthetic reactions. All radical SAM enzymes contain one or more iron‐sulphur clusters, with SAM binding directly to an iron‐sulphur cluster in a manner that facilitates reduction of SAM and generation of a highly reactive 5′‐deoxyadenosyl radical intermediate. This potent intermediate typically initiates the subsequent radical transformations by abstracting a hydrogen atom from the substrate or from a nearby protein residue. Despite the diverse array of reactions catalysed, members of the Radical SAM superfamily share a similar structural topology that facilitates this interaction between SAM and an iron‐sulphur cluster and allows restricted access of substrates to the site of radical generation.

Key Concepts:

  • The Radical SAM superfamily comprises a diverse array of enzymes that contain a common core structural fold and use S‐adenosylmethionine to generate organic radicals.

  • Radical SAM enzymes contain one or more iron‐sulphur clusters that are essential for catalysis.

  • S‐Adenosylmethionine is reduced and cleaved at the sulfonium centre to generate a 5′‐deoxyadenosyl radical.

  • Radical SAM enzymes typically use a 5′‐deoxyadenosyl radical to abstract a hydrogen atom from the substrate.

  • Reaction types catalysed by radical SAM enzymes include oxidation, reduction, methylation, methylthiolation, sulfurylation and complex rearrangement reactions.

  • Radical SAM enzymes are use to introduce several posttranslational and posttranscriptional modifications into proteins and RNA.

  • Radical SAM enzymes are involved in thousands of uncharacterised bacterial natural product biosynthesis pathways.

Keywords: adenosylmethionine; iron‐sulphur cluster; enzyme mechanism; protein structure; radical

Figure 1.

(a) A cartoon depicting the conserved loop that contains the radical SAM consensus sequence, taken from the structure of biotin synthase. The orange tube is the protein backbone, green residues are the three conserved cysteines, blue residue is the semiconserved aromatic residue, all shown relative to the [4Fe‐4S] cluster and SAM. (b) A partial alignment of the conserved [4Fe‐4S]2+/+ cluster coordination site from a selection of radical SAM enzymes. The sequence alignment shows three strictly conserved cysteine residues that coordinate to the FeS cluster (green) and a semiconserved aromatic residue that stacks against the SAM adenine ring (blue). The nonhomologous cluster binding site from the radical SAM enzyme ThiC is also shown for comparison.

Figure 2.

The chemical reactions catalysed by a selection of radical SAM enzymes. (a) Most of the radical SAM enzymes catalyse reduction of the SAM sulfonium with an electron derived from a [4Fe‐4S]+ cluster and a hydrogen atom abstracted from the substrate, resulting in formation of a high‐energy substrate radical. (b–s) The overall chemical transformations catalysed by a selection of radical SAM enzymes. For simplicity, other essential components of several of the reactions are not shown.

Figure 3.

Radical SAM enzymes share a common structural architecture. Shown are ribbon depictions of the structures of monomer subunits of Escherichia coli pyruvate formate‐lyase activating enzyme, E. coli coproporphyrinogen III oxidase (HemN), E. coli biotin synthase (BioB), Thermotoga maritima S12 protein aspartyl‐89 3‐methylthioltransferase (RimO). At the upper region of each protein is the SAM‐[4Fe‐4S]2+ complex coordinate to 3 conserved cysteine residues within an extended loop. The core structure has an (αβ)6 topology generating a six‐stranded parallel β sheet (yellow ribbons) that forms a 3/4‐barrel around the active site. PFL AE has an open partial barrel that presumably accommodates the large protein substrate PFL; a peptide substrate found in the crystal structure is not shown. HemN has a more complete barrel with an empty cleft below SAM that likely binds coproporphyrinogen III. BioB has a complete (αβ)8 barrel that encapsulates the substrate dethiobiotin and the sulphur‐donating [2Fe‐2S]2+ cluster. RimO has a TRAM domain (green) that is thought to interact with the protein substrate or 23S rRNA, and another domain (teal and yellow) that binds a second [4Fe‐4S]2+ cluster. Sulphide (yellow ball) and methanethiol likely bind to this cluster; SAM was not observed in the structure but likely binds to the upper [4Fe‐4S]2+ cluster. Structures are drawn with PyMol using PDB files 3CB8, 1OLT, 1R30 and 4JC0.

Figure 4.

The complex between SAM and the [4Fe‐4S]2+/+ cluster. (a) The interaction of SAM with the FeS cluster is taken from crystal structure of PFL AE. The carboxylate and amine of the methionyl substituents are coordinated to a unique Fe from the cluster, and together with hydrogen bonds from protein residues to the ribose and adenine (not shown); this interaction forces close proximity of the positively‐charged sulfonium and the FeS cluster. Two views show the alignment of SAM with atoms in the FeS cluster. (b) Quantum mechanical calculations show that the highest energy electron in the reduced cluster is delocalised in a large molecular orbital (highest energy occupied molecular orbital, HOMO) that includes all of the [4Fe‐4S]+ cluster atoms and cysteine thiolate ligands, but that does not extend significantly onto SAM. The lowest energy unoccupied molecular orbital (LUMO) is the SAM sulfonium σ* orbital, which includes contributions from atoms in the [4Fe‐4S]+ cluster. Reproduced with permission from Kamachi et al. (). © Elsevier. (c) A mechanism for SAM sulfonium cleavage in which the reduced [4Fe‐4S]+ cluster contains a high‐energy electron delocalised throughout the Fe and S atoms. Promotion of this electron to the LUMO results in the electron being delocalised onto the sulfonium, resulting in a weakened three‐electron C–S bond, which likely represents the transition state for bond cleavage. Following bond cleavage, methionine forms a stable complex with the [4Fe‐4S]2+ cluster, and the now planar 5′‐dA• methylene radical can react with substrate on the face opposite the cluster.



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Further Reading

Atta M, Mulliez E, Arragain S et al. (2010) S‐Adenosylmethionine‐dependent radical‐based modification of biological macromolecules. Current Opinion in Structural Biology 20: 684–692.

Bandarian V (2012) Radical SAM enzymes involved in the biosynthesis of purine‐based natural products. Biochimica et Biophysica Acta 1824: 1245–1253.

Booker SJ and Grove TL (2010) Mechanistic and functional versatility of radical SAM enzymes. Faculty of 1000 Biology Reports 2: 52.

Challand MR, Driesener RC and Roach PL (2011) Radical S‐adenosylmethionine enzymes: mechanism, control and function. Natural Products Reports 28: 1696–1721.

Dowling DP, Vey JL, Croft AK and Drennan CL (2012) Structural diversity in the AdoMet radical enzyme superfamily. Biochimica et Biophysica Acta 1824: 1178–1195.

Duffus BR, Hamilton TL, Shepard EM et al. (2012) Radical AdoMet enzymes in complex metal cluster biosynthesis. Biochimica et Biophysica Acta 1824: 1254–1263.

Frey PA, Hegeman AD and Ruzicka FJ (2008) The Radical SAM Superfamily. Critical Reviews of Biochemistry and Molecular Biology 43: 63–88.

Lanz ND and Booker SJ (2012) Identification and function of auxiliary iron‐sulphur clusters in radical SAM enzymes. Biochimica et Biophysica Acta 1824: 1196–1212.

Ruszczycky MW, Ogasawara Y and Liu HW (2012) Radical SAM enzymes in the biosynthesis of sugar‐containing natural products. Biochimica et Biophysica Acta 1824: 1231–1244.

Shisler KA and Broderick JB (2012) Emerging themes in radical SAM chemistry. Current Opinion in Structural Biology 22: 701–710.

Zhang Q and Liu W (2011) Complex biotransformations catalysed by radical S‐adenosylmethionine enzymes. Journal of Biological Chemistry 286: 30245–30252.

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Jarrett, Joseph T, and Farrar, Christine E(Nov 2014) Radical S‐Adenosylmethionine (SAM) Superfamily. In: eLS. John Wiley & Sons Ltd, Chichester. http://www.els.net [doi: 10.1002/9780470015902.a0020547.pub2]