We intend to probe enzymatic reaction mechanisms by two main lines of approach, the first with mechanism-based enzyme inactivators, the second via study of fluorinated substrate analogs. Mechanism-Based Inactivators or Suicide Substrates require catalytic unmasking of a latent group by the enzyme at its active site, leading to covalent inactivation. The approach provides mechanistic information, structural information on active site catalytic components, and has real in vivo utility. We will investigate allylsulfoxide inactivators of amine oxidases, flavoprotein S-oxygenases, and live cytochrome P450 monooxygenases. We will also study the mechanism of action of neuroconvulsive allylglycine on glutamate decarboxylase, the lathyritic agent beta-aminopropionitrile on elastin lysyl oxidase, and fluorinated amino acid lactones on serine hydroxymethylase. Fluorinated substrates involve all four separate diastereomers of 2-fluorocitrate on various citrate-utilizing enzymes and also the preparation and utilization of chiral 2-(3H)-2-fluoroacetylCoA with claisen condensation enzymes (citrate synthase , malate synthase). Also 3-(3H)-3-fluoropyruvate will be prepared in chiral form and used with biotin-dependent carboxylases and thiamin-dependent decarboxylases to probe enzyme stereochemistry. In the fluorinated substrate work, enzyme stereospecificity, or lack thereof, is linked to molecular bases of toxicity.
Showing the most recent 10 out of 58 publications
