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Literature summary for 2.8.1.12 extracted from
- Mechanistic and mutational studies of Escherichia coli molybdopterin synthase clarify the final step of molybdopterin biosynthesis (2003), J. Biol. Chem., 278, 14523-14532.
Cloned(Commentary)
| Cloned (Comment) | Organism |
|---|---|
| genes moaD and moaE, expression of intein-fusion thiocarboxylated MoaD, expression of wild-type and mutant MoaDs and MoaEs in Escherichia coli strain BL21(DE3) | Escherichia coli |
Protein Variants
| Protein Variants | Comment | Organism |
|---|---|---|
| E128K | site-directed mutagenesis, mutant of MoaE, the MPT synthase containing E128K MoaE is 16.6times slower than the wild-type protein | Escherichia coli |
| E141DELTA | site-directed mutagenesis, the mutant shows reduced activity compared to the wild-type enzyme, 12.3times slower than the wild-type protein, truncation of the C-terminal helix might be predicted to disrupt interaction with MoaD-SH | Escherichia coli |
| F34A | site-directed mutagenesis, mutant of MoaE, the mutant shows reduced activity compared to the wild-type enzyme | Escherichia coli |
| G81DELTA | deletion of the MoaD C-terminal glycine (G81DELTA MoaD-SH) completely abolishes MPT synthase activity | Escherichia coli |
| K119A | site-directed mutagenesis, mutant of MoaE, the mutant shows loss of activity, probably due to complete failure to form the heterotetramer | Escherichia coli |
| K126A | site-directed mutagenesis, mutant of MoaE, the mutant shows reduced activity compared to the wild-type enzyme. The intermediate, rather than MPT, is the major product of the K126A synthase reaction | Escherichia coli |
| M115A | site-directed mutagenesis, mutant of MoaE, the mutant shows reduced activity compared to the wild-type enzyme | Escherichia coli |
| R140A | site-directed mutagenesis, mutant of MoaE, the mutant shows reduced activity compared to the wild-type enzyme | Escherichia coli |
| R39A | site-directed mutagenesis, mutant of MoaE, the mutant shows reduced activity compared to the wild-type enzyme | Escherichia coli |
Natural Substrates/ Products (Substrates)
| Natural Substrates | Organism | Comment (Nat. Sub.) | Natural Products | Comment (Nat. Pro.) | Rev. | Reac. |
|---|---|---|---|---|---|---|
| cyclic pyranopterin phosphate + 2 [molybdopterin-synthase sulfur-carrier protein]-Gly-NH-CH2-C(O)SH + H2O | Escherichia coli | - |
molybdopterin + 2 molybdopterin-synthase sulfur-carrier protein | - |
? |  |
Organism
| Organism | UniProt | Comment | Textmining |
|---|---|---|---|
| Escherichia coli | - |
- |
- |
Purification (Commentary)
| Purification (Comment) | Organism |
|---|---|
| recombinant intein-fusion wild-type and mutant MoaDs and MoaEs from Escherichia coli strain BL21(DE3) by ammonium sulfate fractionation, gel filtration, and chitin affinity chromatography, intein cleavage, followed by another step of gel filtration | Escherichia coli |
Reaction
| Reaction | Comment | Organism | Reaction ID |
|---|---|---|---|
| cyclic pyranopterin phosphate + 2 [molybdopterin-synthase sulfur-carrier protein]-Gly-NH-CH2-C(O)SH + H2O = molybdopterin + 2 molybdopterin-synthase sulfur-carrier protein | reaction mechanism via an intermediate, that remains tightly associated with the protein, overview. Stoichiometry of 2 molecules of thiocarboxylated MoaD per conversion of a single precursor Z molecule to molybdopterin, protein-bound intermediate show a monosulfurated structure that contains a terminal phosphate group similar to that present in molybdopterin | Escherichia coli |
Substrates and Products (Substrate)
| Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
|---|---|---|---|---|---|---|
| cyclic pyranopterin phosphate + 2 [molybdopterin-synthase sulfur-carrier protein]-Gly-NH-CH2-C(O)SH + H2O | - |
Escherichia coli | molybdopterin + 2 molybdopterin-synthase sulfur-carrier protein | - |
? | |
Subunits
| Subunits | Comment | Organism |
|---|---|---|
| More | MoaD is capable of forming two different stable, yet reversible, heterotetrameric complexes that perform biochemically distinct reactions involving the C terminus of MoaD | Escherichia coli |
| tetramer | dimer of dimers containing the MoaD and MoaE proteins, the enzyme is an alpha2beta2 heterotetramer of the smaller MoaD and larger MoaE proteins | Escherichia coli |
Synonyms
| Synonyms | Comment | Organism |
|---|---|---|
| MPT synthase | - |
Escherichia coli |
Temperature Optimum [°C]
| Temperature Optimum [°C] | Temperature Optimum Maximum [°C] | Comment | Organism |
|---|---|---|---|
| 22 | - |
assay at room temperature | Escherichia coli |
pH Optimum
| pH Optimum Minimum | pH Optimum Maximum | Comment | Organism |
|---|---|---|---|
| 7.2 | - |
assay at | Escherichia coli |
General Information
| General Information | Comment | Organism |
|---|---|---|
| metabolism | molybdopterin (MPT) synthase catalyzes the final step in the biosynthesis of MPT, the metal-binding organic portion of the molybdenum cofactor, Moco | Escherichia coli |
| additional information | Lys-119 is a residue essential for activity and Arg-39 and Lys-126 are also residues critical for the reaction | Escherichia coli |
| physiological function | the enzyme is involved in the biosynthesis of the molybdenum cofactor. In the cofactor, the metal is complexed to the unique cis-dithiolene moiety located on the pyran ring of molybdopterin, molybdopterin synthase is responsible for adding the dithiolene to a desulfo precursor termed precursor Z. The sulfur used for dithiolene formation is carried in the form of a thiocarboxylate at the MoaD C terminus. The unique cis-dithiolene moiety of MPT chelates molybdenum or tungsten within Moco. The MoeB protein is essential for the formation of the MoaD thiocarboxylate, serving in the transfer of the dithiolene sulfur atoms, is essential for MPT synthase activity | Escherichia coli |
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