Synthesis and kinetic analysis of inhibitors and inactivators of phosphotriesterase Academic Article uri icon

abstract

  • The phosphotriesterase from Pseudomonas diminuta is a highly efficient metalloenzyme which catalyzes the hydrolysis of a variety of organophosphorus nerve agents. The X-ray structure of the enzyme reconstituted with Cd 2+ shows that a coupled binuclear metal center is embedded within a clus ter of histidine residues in the active site. The role of the binuclear metal center in binding and catalysis remains unknown although Bronsted plots have indicated the likelihood for protein-substrate interaction between the metals and the polarizable sites of the substrate. Evaluation of the potential role of the binuclear metal center has now been probed with mimetics of phosphotriesters prepared by substitution of sulfur- nitrogen and carbon for the bridging oxygen of the leaving group. The phosphorothiolate analogues were found to be good substrates while phosphonates bearing monofluoro, difluoro or hydroxyl substituents at the methylene position were found to be the noncompetitive inhibitors with Ki = 0.6-5.5 mM versus paraoxon. Phosphoroamidates were not inhibitors of the enzyme. Bromovinyl and dibromoethyl phosphotriesters were synthesized and tested for their efficiency as mechanism-based inhibitors of the enzyme. 1-Bromovinyl phosphate was highly reactive, resulting in covalent inactivation of the enzyme.

published proceedings

  • FASEB Journal

author list (cited authors)

  • Hong, S. B., Mullins, L., & Raushel, F.

complete list of authors

  • Hong, SB||Mullins, L||Raushel, F

publication date

  • December 1996