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Synthesis and evaluation of new alpha-halovinyl amino acids as amino acid decarboxylase inactivators

Wan Jin Jahng, University of Nebraska - Lincoln


From the medicinal point of view, several amino acid decarboxylases (Arg DC, DOPA DC, Glu DC, His DC, Lys DC, Orn DC, SAM DC) are important targets for the development of specific and potent inactivators. Higher amino acid analogues, in which the α-hydrogen of amino acid is replaced by a chlorovinyl or fluorovinyl group, are potential mechanism-based enzyme inhibitors for their cognate amino acid decarboxylases. This dissertation describes, to our knowledge, the first syntheses and enzymatic evaluation of these two families of quaternary amino acids. ^ Our synthetic approach to quaternary α-chloro and α-bromovinyl amino acids is based upon addition of the appropriate phenylselenenyl halide to suitably protected α-vinyl amino acids, followed by selenide oxidation and pyrolysis. Under conditions of kinetic control (PhSeX, MeCN, 2 ∼ 3 hours), primarily the (±)-(E)-α-(2-chlorovinyl) amino acids are obtained. At longer times for the PhSeX addition step, the (±)-α-(1-chlorovinyl) regioisomer is seen, comprising 50 ∼ 100% of the α-halovinyl amino acids mixture. ^ The direct α-(1-chloro) vinylation using 2,2-dichlorovinyl phenyl sulfone was accomplished as a potential synthetic route to (±)-α-(1-chlorovinyl) amino acids. As proof of principle, free (±)-α-(1-chlorovinyl)alanine and (±)-α-(1-chlorovinyl)glutamate have been synthesized using this chemistry. ^ A series of (±)-(Z)-α-(2-fluorovinyl) amino acids was synthesized from α-formyl amino acids via a Horner-Wadsworth-Emmons reaction with very high stereoselectivity. The resultant α-fluorovinyl sulfone functionality was transformed into an α-fluorovinyl stannane group with high stereoselectivity for all quaternary amino acids examined. Protodestannylation and deprotection were accomplished to give six (±)-(Z)-α-(2-fluorovinyl) amino acids (Ala, DOPA, Glu, Lys, Phe, m-Tyr). ^ A coupled continuous UV assay was used to evaluate inhibitory effects of (±)-α-vinyllysine and (±)-α-vinylarginine. Time-dependent inhibition was observed for both compounds. (±)-(E)-α-(2-Chlorovinyl)lysine, (±)-α-vinyllysine, (±)-(Z)-α-(2-fluorovinyl)lysine, (±)-α-vinylglutamate, and (±)-(Z)-α-(2-fluorovinyl)glutamate were tested with their cognate amino acids decarboxylases with a radioactive time-point assay. Lineweaver-Burk analysis was used to test each assay and to obtain values of Km for each natural substrate (L-lysine/LDC Km = 1.9 mM, L-glutamate/GAD Km = 1.3 mM) and Kitz-Wilson analysis were used to obtain KI, kinact, and t 1/2. Extended dialysis showed the inhibition of lysine decarboxylase to be essentially irreversible. ^ In the case of the best lysine decarboxylase inactivator found, (±)-(Z)-α-(2-fluorovinyl)lysine, non-pseudofirst order kinetics (ln (Et/Eo) vs. preincubation time) were observed. A Kitz-Wilson analysis of these data using tangents to the inactivation curves provided KI = 1.3 mM, kinact = 0.15 min–1 , t1/2 = 5 minutes. Consistent with the notion that this compound is a “suicide substrat”, the product of its enzymatic decarboxylation was observed by 19F NMR and the ratio of turnover product observed to total enzyme inactivated is approximately 46. That the observed 19F NMR signal was due to product has been confirmed by independent synthesis of (Z)-α-(2-fluorovinyl)cadaverine. ^

Subject Area

Chemistry, Organic

Recommended Citation

Jahng, Wan Jin, "Synthesis and evaluation of new alpha-halovinyl amino acids as amino acid decarboxylase inactivators" (2000). ETD collection for University of Nebraska - Lincoln. AAI9976998.