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Asymmetric synthesis of (+)-L-α-(2′Z-fluoro)vinyllysine and its evaluation as inhibitor of lysine decarboxylase

Roberto de la Salud-Bea, University of Nebraska - Lincoln


An asymmetric synthesis of (+)-L-α-(2'Z-fluoro)vinyllysine (L-2FVL) has been achieved. The method is based on the introduction of an electrophilic lysine side chain synthon into the alpha position of vinylglycine-derived dienolate bearing the Comins auxiliary, trans-2-[(2'-β-naphthyl)-2'-propyl]cyclohexanol, as the source of chirality. The L-absolute stereochemistry has been validated by X-ray crystallography (anomalous dispersion on the bis(hydrochloride) salt of the free amino acid). Since both enantiomers of the auxiliary are available, the procedure is also applicable to the synthesis of the (-)-D-enantiomer and, in principle, can be applied to other amino acids using the proper side chain electrophile. Enzymatic studies carried out with L- and D-2FVL as potential inhibitors of lysine decarboxylase (from Hafnia alvei) have demonstrated a substantial difference in activity between the enantiomers. D-2FVL behaves largely, if not exclusively, as a substrate while the L-isomer shows potent time-dependent inactivation of lysine decarboxylase (LDC). Several enzyme kinetic experiments were performed with the L-isomer. Values of KI = 86 ± 22 µM and kinact = 0.26 ± 0.07 min -1 were estimated from a Kitz-Wilson analysis. A partition ratio (total number of turnovers, including "turnovers" leading to inactivation, per inactivation event) of 20 ± 3 was estimated by titration of the enzyme, using two different steady state kinetic assays (spectrophotometric and radioactivity counting) to measure remaining enzyme activity. A value of 16 ± 2 was obtained in complementary experiments using 19F NMR to determine the formation/consumption of fluorinated species during the course of enzyme inactivation reaction. Reversibility of inhibition was examined by exhaustive dialysis of LDC, following inactivation by L-2FVL. At most 12% of the enzyme activity could be restored, but only if the dialysis buffer contained 100 µM PLP. Substrate protection experiments with L-lysine indicate that inactivation is an active site-directed process and that L-2FVL competes well with L-lysine. For instance, 15 min incubation of LDC with 0.25 mM L-2FVL results in 72% inactivation. However, in the presence of 0.25 mM L-lysine only 34% inactivation is observed under the same conditions. In conclusion, we can support L-2FVL as a mechanism-based inactivator of LDC since all the experiments are in accordance with the criteria for this type of trigger.

Subject Area

Organic chemistry

Recommended Citation

Salud-Bea, Roberto de la, "Asymmetric synthesis of (+)-L-α-(2′Z-fluoro)vinyllysine and its evaluation as inhibitor of lysine decarboxylase" (2006). ETD collection for University of Nebraska - Lincoln. AAI3237598.