7-carboxy-7-deazaguanine synthase
This enzyme is involved in the pathway of 7-cyano-7-deazaquanine (preQ0) biosynthesis and catalyses the complex heterocyclic radical-mediated conversion of 6-carboxy-5,6,7,8-tetrahydropterin (CPH4) to 7- carboxy-7-deazaguanine (CDG). Although this step is common to the biosynthetic pathways of all 7-deazapurine-containing compounds the radical SAM enzyme QueE itself is quite variable. This entry represents a variant form in which the three-Cys motif that binds the signature 4Fe-4S cluster takes the form Cx14CxxC, as in Burkholderia multivorans ATCC 17616. The 3D structure of this form has been solved and the mechanism of other QueE proteins is thought to be broadly similar to the one proposed here.
Reference Protein and Structure
- Sequence
-
A0A0H3KB22
(4.3.99.3)
(Sequence Homologues) (PDB Homologues)
- Biological species
-
Burkholderia multivorans ATCC 17616 (Bacteria)
- PDB
-
4njk
- Crystal Structure of QueE from Burkholderia multivorans in complex with AdoMet, 7-carboxy-7-deazaguanine, and Mg2+
(1.911 Å)
- Catalytic CATH Domains
-
3.20.20.70
(see all for 4njk)
- Cofactors
- Magnesium(2+) (1), Tetra-mu3-sulfido-tetrairon (1), S-adenosyl-l-methionine (1)
Enzyme Reaction (EC:4.3.99.3)
Enzyme Mechanism
Introduction
The C5′ of the dA moiety of AdoMet is close (3.9 Å) to the substrate C6, which is poised to abstract the C6 hydrogen after reductive cleavage of AdoMet. Mg(II) may facilitate this H-atom abstraction through its interaction with both the carbonyl and carboxylate moieties of the substrate. After substrate radical formation, a number of routes are possible to generate the five-membered pyrrole ring, all of which are reminiscent of group migration reactions that are catalysed by adenosylcobalamin-dependent and AdoMet radical mutases.
By analogy to lysine aminomutases, one may envision a migration through an azocyclopropyl radical intermediate that is stabilised by resonance. Alternatively, the rearrangement may proceed by ring opening followed by a 5-exo-trig ring closure, resulting in the pyrroline ring. Next, the amino-centred radical would re-abstract a hydrogen atom from dA-H to reform AdoMet for use in a subsequent catalytic cycle, forming a gem-amino carboxylate pyrrole-like ring. The next step, elimination of the amino group must be enzyme catalysed based on the stereoselective loss of deuterium from the substrate C7 position. Here, enzyme-bound Mg(II) could act as a Lewis acid; alternatively, the protein C-terminal carboxylic acid group could promote the reaction through its interaction with the six-membered ring of substrate. Subsequent deprotonation of the pro-R C8 proton of either intermediate would lead to the final aromatised product.
Catalytic Residues Roles
UniProt | PDB* (4njk) | ||
Asp50 | Asp50(70)A | Activates metal binding threonine | activator |
Thr51 | Thr51(71)A | Binds Mg(II) ion | metal ligand |
Cys49, Cys31, Cys46 | Cys49(69)A, Cys31(51)A, Cys46(66)A | Binds [4Fe-4S]-AdoMet cluster | metal ligand |
Glu116 | Glu116(136)A | General acid/base | proton acceptor, proton donor |
Phe25, His204 | Phe25(45)A, His204(224)A | Binds substrate | electrostatic stabiliser |
Chemical Components
intramolecular homolytic elimination, electron transfer, intermediate formation, cofactor used, hydrogen transfer, cyclisation, intramolecular rearrangement, decyclisation, proton transfer, overall product formed, homolysis, inferred reaction step, native state of cofactor regenerated, native state of enzyme regeneratedReferences
- Dowling DP et al. (2014), Nat Chem Biol, 10, 106-112. Radical SAM enzyme QueE defines a new minimal core fold and metal-dependent mechanism. DOI:10.1038/nchembio.1426. PMID:24362703.
- Jäger CM et al. (2017), Chemistry, 23, 953-962. Radical Reaction Control in the AdoMet Radical Enzyme CDG Synthase (QueE): Consolidate, Destabilize, Accelerate. DOI:10.1002/chem.201604719. PMID:27859789.
- Bruender NA et al. (2017), J Am Chem Soc, 139, 1912-1920. 7-Carboxy-7-deazaguanine Synthase: A Radical S-Adenosyl-l-methionine Enzyme with Polar Tendencies. DOI:10.1021/jacs.6b11381. PMID:28045519.
- Bandarian V et al. (2015), Curr Opin Struct Biol, 35, 116-124. Radical-mediated ring contraction in the biosynthesis of 7-deazapurines. DOI:10.1016/j.sbi.2015.11.005. PMID:26643180.
Step 1. Activation step in which the 5'C-S bond in SAM is reductively cleaved via the iron-sulfur cluster (which has been reduced by a single electron from an external protein source), generating the catalytic 5'-deoxyadenosyl radical.
Download: Image, Marvin FileCatalytic Residues Roles
Residue | Roles |
---|---|
Cys31(51)A | metal ligand |
Cys49(69)A | metal ligand |
Cys46(66)A | metal ligand |
Thr51(71)A | metal ligand |
Asp50(70)A | activator |
Phe25(45)A | electrostatic stabiliser |
His204(224)A | electrostatic stabiliser |
Chemical Components
ingold: intramolecular homolytic elimination, electron transfer, intermediate formation, cofactor usedStep 2. The 5--deoxyandenosyl radical abstracts a hydrogen atom from the substrate.
Download: Image, Marvin FileCatalytic Residues Roles
Residue | Roles |
---|---|
Phe25(45)A | electrostatic stabiliser |
His204(224)A | electrostatic stabiliser |
Cys31(51)A | metal ligand |
Cys49(69)A | metal ligand |
Cys46(66)A | metal ligand |
Thr51(71)A | metal ligand |
Chemical Components
hydrogen transferStep 3. The radical intermediate undergoes intramolecular rearrangement.
Download: Image, Marvin FileCatalytic Residues Roles
Residue | Roles |
---|---|
Cys31(51)A | metal ligand |
Cys49(69)A | metal ligand |
Cys46(66)A | metal ligand |
Thr51(71)A | metal ligand |
Phe25(45)A | electrostatic stabiliser |
His204(224)A | electrostatic stabiliser |
Chemical Components
cyclisation, intramolecular rearrangementCatalytic Residues Roles
Residue | Roles |
---|---|
Phe25(45)A | electrostatic stabiliser |
His204(224)A | electrostatic stabiliser |
Asp50(70)A | activator |
Cys31(51)A | metal ligand |
Cys49(69)A | metal ligand |
Cys46(66)A | metal ligand |
Thr51(71)A | metal ligand |
Chemical Components
intramolecular rearrangement, decyclisationStep 5. The amine radical of the intermediate abstracts a hydrogen from the 5'-adenosine group.
Download: Image, Marvin FileCatalytic Residues Roles
Residue | Roles |
---|---|
Cys31(51)A | metal ligand |
Cys49(69)A | metal ligand |
Cys46(66)A | metal ligand |
Thr51(71)A | metal ligand |
Phe25(45)A | electrostatic stabiliser |
His204(224)A | electrostatic stabiliser |
Asp50(70)A | activator |
Chemical Components
hydrogen transferStep 6. The intermediate undergoes a double bond rearrangement to form the reactive intermediate for the ammonia elimination.
Download: Image, Marvin FileCatalytic Residues Roles
Residue | Roles |
---|---|
Cys31(51)A | metal ligand |
Cys49(69)A | metal ligand |
Cys46(66)A | metal ligand |
Thr51(71)A | metal ligand |
Phe25(45)A | electrostatic stabiliser |
His204(224)A | electrostatic stabiliser |
Asp50(70)A | activator |
Chemical Components
intramolecular rearrangementStep 7. The oxyanion collapses to eliminate ammonia with concomitant deprotonation of one of the water ligands of the magnesium ion.
Download: Image, Marvin FileCatalytic Residues Roles
Residue | Roles |
---|---|
Cys31(51)A | metal ligand |
Cys49(69)A | metal ligand |
Cys46(66)A | metal ligand |
Thr51(71)A | metal ligand |
Phe25(45)A | electrostatic stabiliser |
His204(224)A | electrostatic stabiliser |
Asp50(70)A | activator |
Chemical Components
proton transfer, intramolecular rearrangement, overall product formedStep 8. Another double bond rearrangement occurs to set up the next step.
Download: Image, Marvin FileCatalytic Residues Roles
Residue | Roles |
---|---|
Phe25(45)A | electrostatic stabiliser |
His204(224)A | electrostatic stabiliser |
Cys31(51)A | metal ligand |
Cys49(69)A | metal ligand |
Cys46(66)A | metal ligand |
Thr51(71)A | metal ligand |
Asp50(70)A | activator |
Chemical Components
intramolecular rearrangementStep 9. Glu116 abstracts a proton from the intermediate to generate the final product.
Download: Image, Marvin FileCatalytic Residues Roles
Residue | Roles |
---|---|
Cys31(51)A | metal ligand |
Cys49(69)A | metal ligand |
Cys46(66)A | metal ligand |
Thr51(71)A | metal ligand |
Phe25(45)A | electrostatic stabiliser |
His204(224)A | electrostatic stabiliser |
Asp50(70)A | activator |
Glu116(136)A | proton acceptor |
Chemical Components
proton transfer, intramolecular rearrangement, overall product formedStep 10. The enzyme active site is regenerated. The SAM cofactor is reformed and Glu116 donates its proton to a water molecule in an inferred return step.
Download: Image, Marvin FileCatalytic Residues Roles
Residue | Roles |
---|---|
Cys31(51)A | metal ligand |
Cys49(69)A | metal ligand |
Cys46(66)A | metal ligand |
Thr51(71)A | metal ligand |
Asp50(70)A | activator |
Glu116(136)A | proton donor |