Solvent oxygen is not incorporated into N10-formyltetrahydrofolate in the reaction catalyzed by N10-formyltetrahydrofolate synthetase.
The mechanism of the reaction catalyzed by N10-formyltetrahydrofolate synthetase involves the formation of formyl phosphate as an intermediate which then formylates tetrahydrofolate at the N-10 position. Previous studies demonstrated that the non-enzymic formylation of tetrahydrofolate by formyl phosphate occurs exclusively at the more nucleophilic 5-nitrogen in the reduced pyrazine ring. The experiments described in this report were designed to determine whether N5-formyltetrahydrofolate might be the first product to be formed on the enzyme, followed by formyl transfer to the 10-nitrogen via the cyclic intermediate N5,10-methenyltetrahydrofolate. If this were the case, oxygen from solvent H2O would be incorporated into the formyl group of the N10-derivative. By conducting the reaction in a 1:1 mixture of [16O]H2O and [18O]H2O and using 13C NMR spectroscopy we show that no 18O is incorporated into the product and conclude that the reaction proceeds via a direct formylation of the N-10 position by formyl phosphate.
Song
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Department of Biochemistry, University of Kansas, Lawrence 66045-2106.
Jahansouz
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Himes
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RH
D002384:Catalysis; D003013:Clostridium / Q000201:enzymology; D005574:Formate-Tetrahydrofolate Ligase / Q000737:chemistry*; D005575:Formyltetrahydrofolates / Q000737:chemistry* / Q000378:metabolism; D009682:Magnetic Resonance Spectroscopy; D010100:Oxygen / Q000737:chemistry*; D012997:Solvents
D005575:Formyltetrahydrofolates; D012997:Solvents; D005574:Formate-Tetrahydrofolate Ligase; D010100:Oxygen