Uroporphyrinogen-III C-methyltransferase

 

The reference protein (CysG) covers three different domains/functions each component of which can be found as independent proteins in other organisms. This entry relates to the uroporphyrinogen-III C-methyltransferase function (residues 216-448 in the reference protein). This catalytic site catalyses two sequential methylation reactions using S-andenosyl-L-methionine (SAM), the first forming precorrin-1 and the second leading to the formation of precorrin-2. It is the first of three steps leading to the formation of siroheme from uroporphyrinogen III.

 

Reference Protein and Structure

Sequence
P25924 UniProt (1.3.1.76, 2.1.1.107, 4.99.1.4) IPR006366 (Sequence Homologues) (PDB Homologues)
Biological species
Salmonella enterica subsp. enterica serovar Typhimurium str. LT2 (Bacteria) Uniprot
PDB
1pjq - Structure and function of CysG, the multifunctional methyltransferase/dehydrogenase/ferrochelatase for siroheme synthesis (2.21 Å) PDBe PDBsum 1pjq
Catalytic CATH Domains
3.40.1010.10 CATHdb 3.30.950.10 CATHdb (see all for 1pjq)
Click To Show Structure

Enzyme Reaction (EC:2.1.1.107)

uroporphyrinogen III(8-)
CHEBI:57308ChEBI
+
S-adenosyl-L-methionine zwitterion
CHEBI:59789ChEBI
→
hydron
CHEBI:15378ChEBI
+
S-adenosyl-L-homocysteine zwitterion
CHEBI:57856ChEBI
+
precorrin-2(7-)
CHEBI:58827ChEBI
Alternative enzyme names: S-adenosyl-L-methionine-dependent uroporphyrinogen III methylase, CobA, CysG, SUMT, SirA, Adenosylmethionine-uroporphyrinogen III methyltransferase, Uroporphyrinogen methyltransferase, Uroporphyrinogen-III methylase, Uroporphyrinogen III methylase, Urogen III methylase, Uroporphyrinogen-III methyltransferase, Uroporphyrin-III C-methyltransferase, S-adenosyl-L-methionine:uroporphyrin-III C-methyltransferase,

Enzyme Mechanism

Introduction

The methyl transfers occur through SN2 type reactions in which the incoming methyl group undergoes stereochemical inversion. The nitrogen lone pair of ring A activates carbon C2 which subsequently performs nucleophilic attack on the methyl group bound to the sulfur of SAM. Asp248 carboxylate group abstracts a proton from the tetrapyrrole ring. Lys270 could serve as a general acid to promote tautomerization of the tetrapyrrole, facilitating proton abstraction by Asp248. The resultant rearrangement of double bonds (C1-NH+ and C3-C4 to C20-C1 and C4-C5 respectively) forms precorrin-1. The resultant S-adenosyl-L-homocystine (SAH) and precorrin-1 are lost from the active site followed by binding of new SAM and precorrin-1 in a new orientation placing C7 in the correct position for methylation. The above process is repeated at C7, forming a second SAH and precorrin-2.

Catalytic Residues Roles

UniProt PDB* (1pjq)
Asp248 Asp248A Abstracts a proton from the substrate to cause double bond rearrangement forming precorrin-1. Then abstracts a proton from the substrate to cause double bond rearrangement forming precorrin-2. proton acceptor, electrostatic stabiliser, proton donor
Lys270 Lys270A Acts as a general acid to aid proton abstraction by Asp248. electrostatic stabiliser
Met382 Met382A May function to destabilize the charged S of SAM thus favouring formation of SAH. electrostatic stabiliser
*PDB label guide - RESx(y)B(C) - RES: Residue Name; x: Residue ID in PDB file; y: Residue ID in PDB sequence if different from PDB file; B: PDB Chain; C: Biological Assembly Chain if different from PDB. If label is "Not Found" it means this residue is not found in the reference PDB.

Chemical Components

bimolecular nucleophilic substitution, overall reactant used, proton transfer, tautomerisation (not keto-enol), intermediate formation, overall product formed, native state of enzyme regenerated

References

  1. Vévodová J et al. (2004), J Mol Biol, 344, 419-433. Structure/Function Studies on a S-Adenosyl-l-methionine-dependent Uroporphyrinogen III C Methyltransferase (SUMT), a Key Regulatory Enzyme of Tetrapyrrole Biosynthesis. DOI:10.1016/j.jmb.2004.09.020. PMID:15522295.
  2. Rehse PH et al. (2005), Acta Crystallogr D Biol Crystallogr, 61, 913-919. Structure of a closed-form uroporphyrinogen-IIIC-methyltransferase fromThermus thermophilus. DOI:10.1107/s0907444905008838. PMID:15983414.
  3. Stroupe ME et al. (2003), Nat Struct Biol, 10, 1064-1073. CysG structure reveals tetrapyrrole-binding features and novel regulation of siroheme biosynthesis. DOI:10.1038/nsb1007. PMID:14595395.

Step 1. A methyl group from SAM is added to C2 of the pyrrole ring in a biomolecular nucleophilic substitution reaction.

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Catalytic Residues Roles

Residue Roles
Met382A electrostatic stabiliser
Asp248A electrostatic stabiliser
Lys270A electrostatic stabiliser

Chemical Components

ingold: bimolecular nucleophilic substitution, overall reactant used

Step 2. Asp 248 accepts a proton from the intermediate, causing tautomerization of the tetrapyrrole.

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Catalytic Residues Roles

Residue Roles
Lys270A electrostatic stabiliser
Asp248A proton acceptor

Chemical Components

proton transfer, tautomerisation (not keto-enol)

Step 3. Further tautomerization of the enamine occurs and precorrin 1 is formed.

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Catalytic Residues Roles

Residue Roles
Asp248A proton donor

Chemical Components

proton transfer

Step 4. A methyl group is added to precorrin-1 by SAM in a bimolecular nucleophilic substitution reaction.

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Catalytic Residues Roles

Residue Roles
Met382A electrostatic stabiliser
Asp248A electrostatic stabiliser
Lys270A electrostatic stabiliser

Chemical Components

ingold: bimolecular nucleophilic substitution, intermediate formation

Step 5. Asp248 deprotonates the intermediate. Subsequent rearrangement of the resulting enamine double bond forms precorrin 2.

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Catalytic Residues Roles

Residue Roles
Asp248A proton acceptor

Chemical Components

proton transfer, overall product formed

Step 6. In an inferred step Asp248 is deprotonated to regenerate the native state of the enzyme.

Download: Image, Marvin File

Catalytic Residues Roles

Residue Roles
Asp248A proton donor

Chemical Components

proton transfer, native state of enzyme regenerated
Download: Image, Marvin File

Step 1. A methyl group from SAM is added to C2 of the pyrrole ring in a biomolecular nucleophilic substitution reaction.

Step 2. Asp 248 accepts a proton from the intermediate, causing tautomerization of the tetrapyrrole.

Step 3. Further tautomerization of the enamine occurs and precorrin 1 is formed.

Step 4. A methyl group is added to precorrin-1 by SAM in a bimolecular nucleophilic substitution reaction.

Step 5. Asp248 deprotonates the intermediate. Subsequent rearrangement of the resulting enamine double bond forms precorrin 2.

Step 6. In an inferred step Asp248 is deprotonated to regenerate the native state of the enzyme.

Products.

Contributors

Gemma L. Holliday, Amelia Brasnett