Representative enzymes will be mentioned for each class. Radical SAM enzymes and their mechanisms known before 2008 are summarized by Frey
et al. • Radical S-Adenosylmethionine Enzymes: • Molecular architectures and functions of radical enzymes and their proteins: • Radical SAM enzymes in
RiPP biosynthesis. • Radical SAM enzymes with a vitamin B12 (cobalamin)-binding domain.
Carbon methylation Radical SAM
methylases/methyltransferases are one of the largest yet diverse subgroups and are capable of methylating a broad range of unreactive carbon and phosphorus centers. These enzymes are divided into three classes (Class A, B and C) with representative methylation mechanisms. The shared characteristic is the usage of SAM, split into two distinct roles: one as a source of a
methyl group donor, and the second as a source of 5'-dAdo radical. Another class has been proposed (class D) but proved to be wrongly assigned.
Class A sub-family • Class A enzymes methylate specific
adenosine residues on
rRNA and/or tRNA. In other words, they are RNA base-modifying radical SAM enzymes. • The most mechanistically well-characterized are enzymes RlmN and Cfr. Both enzymes methylates substrate by adding a methylene fragment originating from SAM molecule. Therefore, RlmN and Cfr are considered methyl synthases instead of methyltransferases. File:Radical SAM Enzyme Mmp10.tif|thumb|Structure of a B12-dependent radical SAM enzyme (PDB:7QBS)
Class B sub-family • Class B enzymes are the largest and most versatile which can methylate a wide range of carbon and phosphorus centers.
Class C sub-family • Class C enzymes are reported to play roles in biosynthesis of complex natural products and secondary metabolites. These enzymes methylate heteroaromatic substrates • These enzymes contain both the radical SAM motif and exhibit striking sequence similarity to
coproporhyrinogen III oxidase (HemN), a radical SAM enzyme involved in
heme biosynthesis • Jaw5 is suggested to be responsible for
cyclopropane modifications.
Methylthiolation of tRNAs Methylthiotransferases belong to a subset of radical SAM enzymes that contain two [4Fe-4S]+ clusters and one radical SAM domain. Methylthiotransferases play a major role in catalyzing methylthiolation on tRNA nucleotides or
anticodons through a redox mechanism.
Thiolation modification is believed to maintain translational efficiency and fidelity. MiaB and RimO are both well-characterized and bacterial prototypes for tRNA-modifying methylthiotransferases • MiaB introduces a methylthio group to the isopentenylated A37 derivatives in the tRNA of
S. Typhimurium and
E. coli by utilizing one SAM molecule to generate 5'-dAdo radical to activate the substrate and a second SAM to donate a sulfur atom to the substrate. • RimO is responsible for post-translational modification of Asp88 of the
ribosomal protein S12 in
E. coli. The crystal structure sheds light on the mechanistic action of RimO. The enzyme catalyzes pentasulfide bridge formation linking two Fe-S clusters to allow for sulfur insertion to the substrate. eMtaB is the designated methylthiotransferase in eukaryotic and archaeal cells. eMtaB catalyzes the methylthiolation of tRNA at position 37 on N6-threonylcarbamoyladenosine. A bacterial homologue of eMtaB, YqeV has been reported and suggested to function similarly to MiaB and RimO.
Anaerobic oxidative decarboxylation • One well-studied example is HemN. HemN or anaerobic
coproporphyrinogen III oxidase is a radical SAM enzyme that catalyzes the oxidative decarboxylation of
coproporphyrinogen III to
protoporphyrinogen IX, an intermediate in heme biosynthesis. Evidence support the idea that HemN utilizes two SAM molecules to mediate radical-mediated hydrogen transfer for the sequential decarboxylation of the two propionate groups of coproporphyrinogen III. •
Hyperthermophilic sulfate-reducing archaen
Archaeoglobus fulgidus enables anaerobic oxidation of long chain
n-alkanes. PflD is reported to be responsible for the capacity of
A. fulgidus to grow on a wide range of unsaturated carbons and fatty acids. A detailed biochemical and mechanistic characterization of PflD is still undergoing but preliminary data suggest PflD may be a radical SAM enzyme.
Protein post-translational modification • Formyl-glycine dependent sulfatases require the critical post-translational modification of an active site cysteine or
serine residue into a Cα-formylglycine. A radical SAM enzyme called anSME These peptides belong to the emerging class of
ribosomally synthesized and post-translationally modified peptides (RiPPs). • PoyD and PoyC in polytheonamide biosynthesis • TbtI in thiomuracin biosynthesis • EpeE (previously called YydG) in biosynthesis • MoaA in
molybdopterin biosynthesis • MqnE and MqnC in
menaquinone biosynthesis • RumMC2 in ruminococcin C biosynthesis
Epimerization Radical SAM
epimerases are responsible for the
regioselective introduction of
D-amino acids into RiPPs. This peptide cytotoxin is naturally produced by uncultivated bacteria that exist as symbionts in a marine sponge. • YydG (EpeE) epimerase modifies two amino acid positions on YydF in Gram-positive
Bacillus subtilis. Despite the remaining unknowns and controversies involving SPL-catalyzed reaction, it is certain that SPL utilizes SAM as a cofactor to generate 5'-dAdo radical to revert SP to two thymine residues. • HydG is a radical SAM responsible for generating
CO and
CN− ligands in the Hydrogenase#.5BFeFe.5D_hydrogenase|[Fe-Fe]-hydrogenase (HydA) in various anaerobic bacteria. •
Viperin is an
interferon-stimulated radical SAM enzyme which converts CTP to ddhCTP (3ʹ-deoxy-3′,4ʹdidehydro-CTP), which is a chain terminator for viral
RdRps and therefore a natural antiviral compound. == Clinical considerations ==