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Literature summary for 2.8.1.12 extracted from

  • Gutzke, G.; Fischer, B.; Mendel, R.R.; Schwarz, G.
    Thiocarboxylation of molybdopterin synthase provides evidence for the mechanism of dithiolene formation in metal-binding pterins (2001), J. Biol. Chem., 276, 36268-36274.
    View publication on PubMed

Cloned(Commentary)

Cloned (Comment) Organism
genes moaD and moaE,expression of His-tagged wild-type and mutant protein in Escherichia coli strain M15 Escherichia coli

Molecular Weight [Da]

Molecular Weight [Da] Molecular Weight Maximum [Da] Comment Organism
6900
-
2 * 6900, MoaD, + 2 * 16500, MoaE, SDS-PAGE, composed of two small MoaD and two large subunits MoaE. Both forms of MoaD, carboxylated and thiocarboxylated, are monomeric and are able to form a heterotetrameric complex after coincubation in equimolar ratios with MoaE, but only the thiocarboxylated MPT synthase complex is able to convert precursor Z in vitro to MPT Escherichia coli
16500
-
2 * 6900, MoaD, + 2 * 16500, MoaE, SDS-PAGE, composed of two small MoaD and two large subunits MoaE. Both forms of MoaD, carboxylated and thiocarboxylated, are monomeric and are able to form a heterotetrameric complex after coincubation in equimolar ratios with MoaE, but only the thiocarboxylated MPT synthase complex is able to convert precursor Z in vitro to MPT Escherichia coli
46100
-
carboxylated MPT synthase, gel filtration Escherichia coli
52800
-
thiocarboxylated MPT synthase, gel filtration 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 His-tagged wild-type and mutant protein from Escherichia coli strain M15 by nickel affinity chromatography and gel filtration. Elution of carboxylated or thiocarboxylated protein is induced by using a cleavage buffer containing 20 mM Tris/HCl, 500 mM NaCl, 0.1 mM EDTA, pH 8.0, with either 30 mM dithiothreitol or 30 mM ammonium sulfide, respectively. The cleavage reaction is performed at 4°C for at least 24 h 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 two-step reaction of MPT synthesis where the dithiolene is generated by two thiocarboxylates derived from a single tetrameric MPT synthase, hypothetical model for the conversion of precursor Z to molybdopterin in the MPT synthase reaction, overview 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
-
?
additional information in vitro generation of carboxylated and thiocarboxylated MoaD, overview Escherichia coli ?
-
?

Subunits

Subunits Comment Organism
? x * 8757.94, recombinant carboxylated MoaD, sequence calculation, x * 8774.03, recombinant thiocarboxylated MoaD, sequence calculation, x * 8757.72, recombinant carboxylated MoaD, mass spectrometry, x * 8774.00, recombinant thiocarboxylated MoaD, mass spectrometry Escherichia coli
heterotetramer 2 * 6900, MoaD, + 2 * 16500, MoaE, SDS-PAGE, composed of two small MoaD and two large subunits MoaE. Both forms of MoaD, carboxylated and thiocarboxylated, are monomeric and are able to form a heterotetrameric complex after coincubation in equimolar ratios with MoaE, but only the thiocarboxylated MPT synthase complex is able to convert precursor Z in vitro to MPT Escherichia coli

General Information

General Information Comment Organism
additional information Both forms of MoaD, carboxylated and thiocarboxylated, are monomeric and are able to form a heterotetrameric complex after coincubation in equimolar ratios with MoaE, but only the thiocarboxylated MPT synthase complex is able to convert precursor Z in vitro to MPT Escherichia coli
physiological function MPT is formed by incorporation of two sulfur atoms into precursor Z, which is catalyzed by MPT synthase. The thiocarboxylation of the C-terminus of MoaD serves as the source of sulfur that is transferred to precursor Z Escherichia coli