Nuclear magnetic resonance studies of formyltetrahydrofolate synthetase interactions with formate and methylammonium ion.
Using nuclear magnetic resonance techniques, we have measured the internuclear distances separating the nucleotide-bound metal from the carbon and hydrogen nuclei of formate as well as the carbon of methylammonium cation when bound to formyltetrahydrofolate synthetase. Measurements were made of the paramagnetic effect on the spin-lattice relaxation rates (1/T1) of 13C and 1H nuclei arising from the replacement of Mg2+ with Mn2+, which binds to the enzyme in the form of a metal-nucleotide complex. Distances from Mn2+ to the formate carbon and proton were found to be 6.3 and 7.4 A, respectively, in the E . ATP . Mn2+ . formate complex and 6.0 and 7.1 A, respectively, in the E . ADP . Mn2+ . formate complex. When tetrahydrofolate was added to the latter complex, the exchange of formate was greatly reduced and became rate limiting for relaxation. These results are consistent with substantial conformational effects produced by the binding of the cofactor. The distance from Mn2+ to the methylammonium carbon in the E . ADP . Mn2+ . CH3NH+3, E . ADP . Mn2+ . formate . CH3NH3+, and E . ADP . Mn2+ . tetrahydrofolate . CH3NH3+ complexes was estimated to be in the range of 7.4-12 A. However, in the E . ADP . Mn2+ formate . tetrahydrofolate . CH3NH3+ complex, the data suggest that exchange of cation contributes significantly to relaxation. These results, combined with other known features of the enzyme, suggest that there may be a monovalent cation site within the active site of the enzyme.
D003013:Clostridium / Q000201:enzymology; D005574:Formate-Tetrahydrofolate Ligase / Q000378:metabolism*; D005561:Formates; D007700:Kinetics; D008025:Ligases / Q000378:metabolism*; D009682:Magnetic Resonance Spectroscopy; D008433:Mathematics; D008744:Methylamines; D011485:Protein Binding; D000644:Quaternary Ammonium Compounds
D005561:Formates; D008744:Methylamines; D000644:Quaternary Ammonium Compounds; C030544:formic acid; C027451:methylamine; D008025:Ligases; D005574:Formate-Tetrahydrofolate Ligase