Skip to main content
NCBI home page
Biochemical Journal logoLink to Biochemical Journal
. 1988 Apr 1;251(1):89–93. doi: 10.1042/bj2510089

Nuclear magnetic resonance relaxation studies of the interaction of ligands with the monomer and tetramer forms of formyltetrahydrofolate synthetase.

C H Yeh 1, D A Hanna 1, G W Everett 1, R H Himes 1
PMCID: PMC1148967  PMID: 3390163

Abstract

Previous work using n.m.r. spectroscopy to investigate the binding between formyltetrahydrofolate synthetase and its ligands was done using the catalytically active tetrameric form of the enzyme. By removal of specific monovalent cations the tetramer dissociates to four identical, catalytically inactive monomers, which are capable of binding nucleotides with affinities similar to those obtained with the tetramer. In the studies reported here, we examined the interaction of metal-nucleotide, formate and monovalent cations with the monomer using n.m.r. relaxation measurements. We were able to demonstrate that formate binds to the monomer. The spin-lattice relaxation rate (1/T1) of the formate carbon in the monomer.M2+.ADP.formate complex is enhanced when Mg2+ is replaced by Mn2+. By assuming that the exchange of formate is not rate-limiting and that tau c of the monomer is the same as that of the tetramer, the distance between the Mn2+ and the formate carbon was calculated and found to be similar in the monomer and tetramer complexes. The spin-lattice relaxation rates of [13C]trimethylammonium ion (an inactive monovalent cation), [13C]methylammonium and [15N]ammonium ions (both active monovalent cations), were measured in the presence of tetramer, MnADP and formate. The relaxation rates of methylammonium and ammonium ions were enhanced under these conditions whereas the relaxation rate of trimethylammonium ion was not. The results indicate that the active monovalent cations bind near the MnADP binding site. A distance from the Mn2+ to the ammonium nitrogen of between 0.5 and 0.6 nm was calculated.

Full text

PDF
89

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Buttlaire D. H. Purification and properties of formyltetrahydrofolate synthetase. Methods Enzymol. 1980;66:585–599. doi: 10.1016/0076-6879(80)66512-4. [DOI] [PubMed] [Google Scholar]
  2. Buttlaire D. H., Reed G. H., Himes R. H. Equilibrium and water proton relaxation rate enhancement properties of formyltetrahydrofolate synthetase-manganous ion-substrate complexes. J Biol Chem. 1975 Jan 10;250(1):254–260. [PubMed] [Google Scholar]
  3. Buttlaire D. H., Reed G. H., Himes R. Electron paramagnetic resonance and water proton relaxation rate studies of formyltetrahydrofolate synthetase-manganous ion complexes. Evidence for involvement of substrates in the promotion of a catalytically competent active site. J Biol Chem. 1975 Jan 10;250(1):261–270. [PubMed] [Google Scholar]
  4. Curthoys N. P., D'Ari Straus L., Rabinowitz J. C. Formyltetrahydrofolate synthetase. Substrate binding to monomeric subunits. Biochemistry. 1972 Feb 1;11(3):345–349. doi: 10.1021/bi00753a006. [DOI] [PubMed] [Google Scholar]
  5. Curthoys N. P., Scott J. M., Rabinowitz J. C. Folate coenzymes of Clostridium acidi-urici. The isolation of (l)-5,10-methenyltetrahydropteroyltriglutamate, its conversion to (l)-tetrahydropteroyltriglutamate and (l)-10-( 14 C)formyltetrahydropteroyltriglutamate, and the synthesis of (l)-10-formyl-(6,7- 3 H 2 )tetrahydropteroyltriglutamate and (l)-(6,7- 3 H 2 )tetrahydropteroyltriglutamate. J Biol Chem. 1972 Apr 10;247(7):1959–1964. [PubMed] [Google Scholar]
  6. Himes R. H., Cohn M. Formyltetrahydrofolate synthetase. Magnetic resonance studies of the reaction. J Biol Chem. 1967 Aug 25;242(16):3628–3635. [PubMed] [Google Scholar]
  7. Himes R. H., Harmony J. A. Formyltetrahydrofolate synthetase. CRC Crit Rev Biochem. 1973 Sep;1(4):501–535. doi: 10.3109/10409237309105441. [DOI] [PubMed] [Google Scholar]
  8. Samuel C. E., D'Ari L., Rabinowitz J. C. Evidence against the folate-mediated formylation of formyl-accepting methionyl transfer ribonucleic acid in Streptococcus faecalis R. J Biol Chem. 1970 Oct 10;245(19):5115–5121. [PubMed] [Google Scholar]
  9. Staben C., Whitehead T. R., Rabinowitz J. C. Heparin-agarose chromatography for the purification of tetrahydrofolate utilizing enzymes: C1-tetrahydrofolate synthase and 10-formyltetrahydrofolate synthetase. Anal Biochem. 1987 Apr;162(1):257–264. doi: 10.1016/0003-2697(87)90035-2. [DOI] [PubMed] [Google Scholar]
  10. Welch W. H., Irwin C. L., Himes R. H. Observations on the monovalent cation requirements of formyltetrahydrofolate synthetase. Biochem Biophys Res Commun. 1968 Feb 15;30(3):255–261. doi: 10.1016/0006-291x(68)90443-9. [DOI] [PubMed] [Google Scholar]
  11. Wendland M. F., Stevens T. H., Buttlaire D. H., Everett G. W., Himes R. H. Nuclear magnetic resonance studies of formyltetrahydrofolate synthetase interactions with formate and methylammonium ion. Biochemistry. 1983 Feb 15;22(4):819–826. doi: 10.1021/bi00273a017. [DOI] [PubMed] [Google Scholar]

Articles from Biochemical Journal are provided here courtesy of The Biochemical Society

RESOURCES