Methane monooxygenase (soluble)
Methyl monooxygenase (MMO) catalyses the conversion of methane to methanol coupled with reduction of oxygen and oxidation of NADH. The soluble type found in Methylosinus trichosporium consists of four components (A to D): protein A (represented by this entry), comprises three chains, in an alpha-2, beta-2, gamma-2 configuration, is a nonheme iron protein containing an unusual mu-hydroxo bridge structure at its active site and interacts with both oxygen and methane.
Methanotrophic bacteria play an essential part in cycling carbon in the biosphere by consuming methane produced in anaerobic sediments and by limiting its flux to the atmosphere where it acts as a greenhouse gas. They have been used as a basis for biomimetic catalysts to convert methane in to the easily transportable liquid form of methanol for use as a fuel.
Reference Protein and Structure
- Sequences
-
P27354
(1.14.13.25)
P27353 (1.14.13.25)
P27355 (1.14.13.25)
(Sequence Homologues)
(PDB Homologues)
- Biological species
-
Methylosinus trichosporium (Bacteria)
- PDB
-
1mhy
- METHANE MONOOXYGENASE HYDROXYLASE
(2.0 Å)
- Catalytic CATH Domains
-
1.10.620.20
(see all for 1mhy)
- Cofactors
- Iron(3+) (2)
Enzyme Mechanism
Introduction
Reduced MMOH reacts with molecular oxygen to form a peroxo intermediate P and oxidising the Fe(II). The Fe(III) ions are further oxidised during the cleavage of the peroxide O-O bond to form another intermediate known as Q. The are a number of possibilities for the mechanism of methane hydroxylation from intermediate Q. In this proposal the reaction begins with approach of the methane to one of the bridging oxo atoms which is accompanied by proton-coupled outer-sphere transfer of the first electron from a C-H bond in methane to one of the Fe(IV) centres. A second electron transfer then occurs to form the methanol product. Both redox reactions are strongly coupled to structural distortions of the diiron core.
The reductase protein MMOR is responsible for accepting electrons from NADH and transferring them to MMOH to reduce the Fe(III) centres back to Fe(II), restoring the active site.
Catalytic Residues Roles
| UniProt | PDB* (1mhy) | ||
| Glu114, His147 | Glu114(109)D(B), His147(142)D(B) | These residues are bound to one of the Fe centres. | metal ligand |
| Glu144, Glu243 | Glu144(139)D(B), Glu243(238)D(B) | These residues are bound to the Fe centres. In the reduced active site they are initially bridging between the two ions, however this changes when molecular oxygen binds. | metal ligand |
| Glu209, His246 | Glu209(204)D(B), His246(241)D(B) | These residues are bound to one of the Fe ions. | metal ligand |
Chemical Components
decoordination from a metal ion, coordination to a metal ion, intermediate formation, redox reaction, electron transfer, homolysis, overall reactant used, overall product formed, radical termination, native state of enzyme regeneratedReferences
- Elango N et al. (1997), Protein Sci, 6, 556-568. Crystal structure of the hydroxylase component of methane monooxygenase from Methylosinus trichosporium OB3b. DOI:10.1002/pro.5560060305. PMID:9070438.
- Han WG et al. (2008), Inorg Chem, 47, 2975-2986. Structural model studies for the peroxo intermediate P and the reaction pathway from P-->Q of methane monooxygenase using broken-symmetry density functional calculations. DOI:10.1021/ic701194b. PMID:18366153.
- Gherman BF et al. (2004), J Am Chem Soc, 126, 2978-2990. Dioxygen activation in methane monooxygenase: a theoretical study. DOI:10.1021/ja036506. PMID:14995216.
- Baik MH et al. (2003), Chem Rev, 103, 2385-2419. Mechanistic studies on the hydroxylation of methane by methane monooxygenase. DOI:10.1021/cr950244f. PMID:12797835.
- Baik M et al. (2002), J Am Chem Soc, 124, 14608-14615. Hydroxylation of Methane by Non-Heme Diiron Enzymes: Molecular Orbital Analysis of C−H Bond Activation by Reactive Intermediate Q. DOI:10.1021/ja026794u.
- Merkx M et al. (2001), Angew Chem Int Ed Engl, 40, 2782-2807. Dioxygen Activation and Methane Hydroxylation by Soluble Methane Monooxygenase: A Tale of Two Irons and Three Proteins A list of abbreviations can be found in Section 7. PMID:11500872.
- Basch H et al. (1999), J Am Chem Soc, 121, 7249-7256. Mechanism of the Methane → Methanol Conversion Reaction Catalyzed by Methane Monooxygenase: A Density Functional Study. DOI:10.1021/ja9906296.
- Nordlund P et al. (1992), FEBS Lett, 307, 257-262. The active site structure of methane monooxygenase is closely related to the binuclear iron center of ribonucleotide reductase. DOI:10.1016/0014-5793(92)80690-i. PMID:1644180.
Step 1. The first step in the reaction involves the departure of the loosely coordinated water molecule trans to the histidines 147 and 246.
Download: Image, Marvin FileCatalytic Residues Roles
| Residue | Roles |
|---|---|
| Glu114(109)D(B) | metal ligand |
| Glu144(139)D(B) | metal ligand |
| His147(142)D(B) | metal ligand |
| Glu209(204)D(B) | metal ligand |
| Glu243(238)D(B) | metal ligand |
| His246(241)D(B) | metal ligand |
Chemical Components
decoordination from a metal ion
Step 2. Molecular oxygen binds to the diiron centre, forming peroxo intermediate P.
Download: Image, Marvin FileCatalytic Residues Roles
| Residue | Roles |
|---|---|
| Glu114(109)D(B) | metal ligand |
| Glu144(139)D(B) | metal ligand |
| His147(142)D(B) | metal ligand |
| Glu209(204)D(B) | metal ligand |
| Glu243(238)D(B) | metal ligand |
| His246(241)D(B) | metal ligand |
Chemical Components
coordination to a metal ion, intermediate formation, redox reaction
Step 3. Glu243 opens up and there is temporary dissociation of Glu144 and the water molecule, allowing for a trans-u-1,2 conformation. The peroxide O-O bond breaks and intermediate Q is formed. The water molecule and Glu144 then rebind to their respective Fe centres. Glu243 forms a hydrogen bond to the water molecule.
Download: Image, Marvin FileCatalytic Residues Roles
| Residue | Roles |
|---|---|
| Glu114(109)D(B) | metal ligand |
| Glu144(139)D(B) | metal ligand |
| His147(142)D(B) | metal ligand |
| Glu209(204)D(B) | metal ligand |
| Glu243(238)D(B) | metal ligand |
| His246(241)D(B) | metal ligand |
| Glu243(238)D(B) | hydrogen bond donor |
Chemical Components
electron transfer, intermediate formation, coordination to a metal ion, decoordination from a metal ion
Step 4. The hydroxylation reaction shown is the Friesner-Lippard model (PMID: ). There is single electron transfer from one bound oxygen to one of the Fe(IV) centres, reducing it to Fe(III). The electrophilicity of the oxo moiety is greatly enhanced by this distortion.
Download: Image, Marvin FileCatalytic Residues Roles
| Residue | Roles |
|---|---|
| Glu114(109)D(B) | metal ligand |
| Glu144(139)D(B) | metal ligand |
| His147(142)D(B) | metal ligand |
| Glu209(204)D(B) | metal ligand |
| Glu243(238)D(B) | metal ligand |
| His246(241)D(B) | metal ligand |
Chemical Components
electron transfer
Step 5. A methane C-H bond is broken by the oxidised oxygen to form a methane radical.
Download: Image, Marvin FileCatalytic Residues Roles
| Residue | Roles |
|---|---|
| Glu114(109)D(B) | metal ligand |
| Glu144(139)D(B) | metal ligand |
| His147(142)D(B) | metal ligand |
| Glu209(204)D(B) | metal ligand |
| Glu243(238)D(B) | metal ligand |
| His246(241)D(B) | metal ligand |
Chemical Components
homolysis, overall reactant used
Step 6. Another electron is transferred from the reduced hydroxyl to the remaining Fe(IV) centre, increasing the electrophilicity of the oxygen moiety.
Download: Image, Marvin FileCatalytic Residues Roles
| Residue | Roles |
|---|---|
| Glu114(109)D(B) | metal ligand |
| Glu144(139)D(B) | metal ligand |
| His147(142)D(B) | metal ligand |
| Glu209(204)D(B) | metal ligand |
| Glu243(238)D(B) | metal ligand |
| His246(241)D(B) | metal ligand |
Chemical Components
redox reaction, electron transfer
Step 7. The newly formed OH group reacts with the methyl radical to give the final product. The active site of the hydroxylase (MMOH) is restored by the reductase protein (MMOR) which also makes up the sMMO enzyme. MMOR contains Fe2S2 and FAD cofactors that enable it to accept electrons from NADH and transfer them to the hydroxylase (not shown here). This reduces the Fe ions to Fe(II).
Download: Image, Marvin FileCatalytic Residues Roles
| Residue | Roles |
|---|---|
| Glu114(109)D(B) | metal ligand |
| Glu144(139)D(B) | metal ligand |
| His147(142)D(B) | metal ligand |
| Glu209(204)D(B) | metal ligand |
| Glu243(238)D(B) | metal ligand |
| His246(241)D(B) | metal ligand |
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