Publications

• Assessment of Pseudomonas aeruginosa N5,N10-methylenetetrahydrofolate dehydrogenase-cyclohydrolase as a potential antibacterial drug target.

  Eadsforth T.C., Gardiner M., Maluf F.V., McElroy S., James D., Frearson J., Gray D., Hunter W.N.

  The bifunctional enzyme methylenetetrahydrofolate dehydrogenase - cyclohydrolase (FolD) is identified as a potential drug target in Gram-negative bacteria, in particular the troublesome Pseudomonas aeruginosa. In order to provide a comprehensive and realistic assessment of the potential of this ta ... >> More

  The bifunctional enzyme methylenetetrahydrofolate dehydrogenase - cyclohydrolase (FolD) is identified as a potential drug target in Gram-negative bacteria, in particular the troublesome Pseudomonas aeruginosa. In order to provide a comprehensive and realistic assessment of the potential of this target for drug discovery we generated a highly efficient recombinant protein production system and purification protocol, characterized the enzyme, carried out screening of two commercial compound libraries by differential scanning fluorimetry, developed a high-throughput enzyme assay and prosecuted a screening campaign against almost 80,000 compounds. The crystal structure of P. aeruginosa FolD was determined at 2.2 Å resolution and provided a template for an assessment of druggability and for modelling of ligand complexes as well as for comparisons with the human enzyme. New FolD inhibitors were identified and characterized but the weak levels of enzyme inhibition suggest that these compounds are not optimal starting points for future development. Furthermore, the close similarity of the bacterial and human enzyme structures suggest that selective inhibition might be difficult to attain. In conclusion, although the preliminary biological data indicates that FolD represents a valuable target for the development of new antibacterial drugs, indeed spurred us to investigate it, our screening results and structural data suggest that this would be a difficult enzyme to target with respect to developing the appropriate lead molecules required to underpin a serious drug discovery effort. << Less

  PLoS ONE 7:E35973-E35973(2012) [PubMed] [EuropePMC]

• 5,10-Methylenetetrahydrofolic dehydrogenase from bakers' yeast. I. Partial purification and some properties.

  RAMASASTRI B.V., BLAKLEY R.L.

  J Biol Chem 237:1982-1988(1962) [PubMed] [EuropePMC]

• The NADP-dependent methylene tetrahydromethanopterin dehydrogenase in Methylobacterium extorquens AM1.

  Vorholt J.A., Chistoserdova L.V., Lidstrom M.E., Thauer R.K.

  An NADP-dependent methylene tetrahydromethanopterin (H4MPT) dehydrogenase has recently been proposed to be involved in formaldehyde oxidation to CO2 in Methylobacterium extorquens AM1. We report here on the purification of this novel enzyme to apparent homogeneity. Via the N-terminal amino acid se ... >> More

  An NADP-dependent methylene tetrahydromethanopterin (H4MPT) dehydrogenase has recently been proposed to be involved in formaldehyde oxidation to CO2 in Methylobacterium extorquens AM1. We report here on the purification of this novel enzyme to apparent homogeneity. Via the N-terminal amino acid sequence, it was identified to be the mtdA gene product. The purified enzyme catalyzed the dehydrogenation of methylene H4MPT with NADP+ rather than with NAD+, with a specific activity of approximately 400 U/mg of protein. It also catalyzed the dehydrogenation of methylene tetrahydrofolate (methylene H4F) with NADP+. With methylene H4F as the substrate, however, the specific activity (26 U/mg) and the catalytic efficiency (Vmax/Km) were approximately 20-fold lower than with methylene H4MPT. Whereas the dehydrogenation of methylene H4MPT (E0 = -390 mV) with NADP+ (E0 = -320 mV) proceeded essentially irreversibly, the dehydrogenation of methylene H4F (E0 = -300 mV) was fully reversible. Comparison of the primary structure of the NADP-dependent dehydrogenase from M. extorquens AM1 with those of methylene H4F dehydrogenases from other bacteria and eucarya and with those of methylene H4MPT dehydrogenases from methanogenic archaea revealed only marginally significant similarity (<15%). << Less

  J. Bacteriol. 180:5351-5356(1998) [PubMed] [EuropePMC]

  This publication is cited by 1 other entry.

• Hydroxymethyl tetrahydrofolic dehydrogenase.

  HATEFI Y., OSBORN M.J., KAY L.D., HUENNEKENS F.M.

  J Biol Chem 227:637-647(1957) [PubMed] [EuropePMC]

• Purification and properties of N5, N10-Methylenetetra-hydrofolate dehydrogenase of calf thymus.

  Yeh Y.C., Greenberg D.M.

  Biochim Biophys Acta 105:279-291(1965) [PubMed] [EuropePMC]