Publication Abstract from PubMed
By using biochemical and structural analyses, we have investigated the catalytic mechanism of the recently discovered flavin-dependent thymidylate synthase ThyX from Paramecium bursaria chlorella virus-1 (PBCV-1). Site-directed mutagenesis experiments have identified several residues implicated in either NADPH oxidation or deprotonation activity of PBCV-1 ThyX. Chemical modification by diethyl pyrocarbonate and mass spectroscopic analyses identified a histidine residue (His53) crucial for NADPH oxidation and located in the vicinity of the redox active N-5 atom of the FAD ring system. Moreover, we observed that the conformation of active site key residues of PBCV-1 ThyX differs from earlier reported ThyX structures, suggesting structural changes during catalysis. Steady-state kinetic analyses support a reaction mechanism where ThyX catalysis proceeds via formation of distinct ternary complexes without formation of a methyl enzyme intermediate.
Catalytic mechanism and structure of viral flavin-dependent thymidylate synthase ThyX.,Graziani S, Bernauer J, Skouloubris S, Graille M, Zhou CZ, Marchand C, Decottignies P, van Tilbeurgh H, Myllykallio H, Liebl U J Biol Chem. 2006 Aug 18;281(33):24048-57. Epub 2006 May 17. PMID:16707489[1]
From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.
