Synthesis Of ?,?-Difluorinated Phosphonate PSer/pThr Mimetics And Their Applications

Serine/threonine phosphorylation is a key protein post-translational modification that regulates the activity of the proteins and impacts diverse cellular signaling conduction pathways.Phosphoproteins and phosphopeptides have shown great importance in the study of phosphorylation's biological processes and protein-protein interactions.However,phosphorylation is unstable in biological system,which brings great challenges to researches because of the existence of phosphatases.Therefore,it is necessary to develop nonhydrolytic phosphorylated amino acids mimics.Phosphatase-inert mimetics,in which nonhydrolyzable methylene?CH₂?or difluoromethylene?CF₂?unit replaces phosphoryl ester oxygen,are widely applied in previous works because they have similar structures with native phosphorylation,and CF₂-substituted phosphorylation mimics are more preferred choices.Nevertheless,the synthetic routes of CF₂-substituted phosphorylation mimics previously reported were either complicated or expensive.Herein we report a novel stereoselective synthetic strategy to construct both CF₂-substituted phosphoserine/phosphothreonine mimetics?Pfa/Pfab?that suitable for solid-phase peptide synthesis?SPPS?,Furthermore we used the former building block to synthesize inhibitors of 14-3-3?.In the synthesis route of Pfa,we started with cheap and commercial available Boc-Ser-OH,which was subsequently converted to iodine substituted alkene.Then it was reacted with CF₂-substituted diethyl phosphonate.Next,the above product was converted to Boc-Pfa?Et?₂-OtBu through asymmetric hydrogenation,then derived for SPPS.This route greatly increased the yield of the target product with which we synthesized the inhibitor of 14-3-3?using SPPS.In the synthesis route of Pfab we started with cheap and available H-Thr-OMe·HCl.The E form halo-alkene was obtained and reacted with CF₂-substituted diethyl phosphonate,Then two chiral centers were constructed through asymmetric hydrogenation in one step to get Ac-Pfab?Et?₂-OMe,which has same conformationwithnative phosphothreonine?theoreticalprediction?.After derivatization we synthesized Fmoc-Pfab-OH.In summary,we have developed a new synthesis route of CF₂-substituted phosphorylated serine/threonine mimics,and applied the former to SPPS.This work is meaningful for the study of phosphoproteins and phosphopeptides.