The influence of fluorine substitution on some enzyme mediated reactions

Abstract

The replacement of a hydrogen or hydroxy group with a fluorine atom is a popular strategy to alter the activity of biologically important molecules, as their similar sizes mean that such a replacement has little steric impact. The effect of fluorine substitution in a number of enzyme mediated processes has been investigated. 3-Fluorocyclohex-l-enylcarbonyl-CoA has been synthesised and the reaction with cyclohexenylcarbonyl-CoA reductase investigated. The fluorinated substrate has a comparable K(_m) value to that of the natural substrate but a V(_max) that is five times greater. A change in the rate-determining step of the reduction was also observed upon fluorine incorporation. The enzyme showed a small but significant stereochemical preference for the production of the axial isomer, consistent with an Anh-Eisenstein model for the transformation. The 6а and 6β isomers of benzyl fluoropenicillanate were synthesised and their methoxide-mediated hydrolyses were investigated. Competitive hydrolysis, using (^19)F NMR spectroscopy, demonstrated that the β isomer was hydrolysed preferentially. A frill kinetic analysis was undertaken, which furnished the rate and equilibrium constants. Monofluorinated enamines were treated in situ with a range a Michael acceptors to afford a variety of novel substituted a-fluoro ketones. 2-Fluorohexanal was synthesised from methyl hexanoate and was demonstrated to be a substrate for the enzyme transketolase with hydroxypyruvate. The enzyme reaction was monitored by (^19)F NMR spectroscopy. The enzyme showed a diastereoselectivity of 9:1 in the condensation of the aldehyde and hydroxypyruvate, and a self-condensation product was also produced. The enzymatic oxidation of the mono- and di-fluoromethylenephosphonate analogues of glycerol-3-phosphate was investigated at neutral pH using a co-factor recycling protocol. The reactions allowed for the first time the identification of the products of oxidation and demonstrated the lability of fluoride via non-enzymatic elimination and stoichiometric defluorination.