Synthesis and Deployment of an Elusive Fluorovinyl Cation Equivalent: Access to Quaternary α-(1′-Fluoro)vinyl Amino Acids as Potential PLP Enzyme Inactivators

Developing specific chemical functionalities to deploy in biological environments for targeted enzyme inactivation lies at the heart of mechanism-based inhibitor development but also is central to other protein-tagging methods in modern chemical biology including activity-based protein profiling and proteolysis-targeting chimeras. We describe here a previously unknown class of potential PLP enzyme inactivators; namely, a family of quaternary, alpha-(1'-fluoro)vinyl amino acids, bearing the side chains of the cognate amino acids. These are obtained by the capture of suitably protected amino acid enolates with beta,beta-difluorovinyl phenyl sulfone, a new (1'-fluoro)vinyl cation equivalent, and an electrophile that previously eluded synthesis, capture and characterization. A significant variety of biologically relevant AA side chains are tolerated including those for alanine, valine, leucine, methionine, lysine, phenylalanine, tyrosine, and tryptophan. Following addition/elimination, the resulting transoid alpha-(1'-fluoro)-beta-(phenylsulfonyl)vinyl AA-esters undergo smooth sulfone-stannane interchange to stereoselectively give the corresponding transoid alpha-(1'-fluoro)-beta-(tributylstannyl)vinyl AA-esters. Protodestannylation and global deprotection then yield these sterically encumbered and densely functionalized quaternary amino acids. The alpha-(1'-fluoro)vinyl trigger, a potential allene-generating functionality originally proposed by Abeles, is now available in a quaternary AA context for the first time. In an initial test of this new inhibitor class, alpha-(1'-fluoro)vinyllysine is seen to act as a time-dependent, irreversible inactivator of lysine decarboxylase from Hafnia alvei. The enantiomers of the inhibitor could be resolved, and each is seen to give time-dependent inactivation with this enzyme. Kitz-Wilson analysis reveals similar inactivation parameters for the two antipodes, L-alpha-(1'-fluoro)vinyllysine (K-i = 630 +/- 20 mu M; t(1/2) = 2.8 min) and D-alpha-(1'-fluoro)vinyllysine (K-i = 470 +/- 30 mu M; t(1/2) = 3.6 min). The stage is now set for exploration of the efficacy of this trigger in other PLP-enzyme active sites.