1.Carboxypeptidase A Inhibition Conscripting Glu-270 And Arg-127: Implications To Carboxypeptidase A Catalytic Actions 2.Arndt-Eistert Reaction Of N-Tosyl Cyclic α-Amino Acids To α, β-Unsaturated Esters Bearing A Terminal Tosylamino Group

Part One. Carboxypeptidase A Inhibition Conscripting Glu-270 and Arg-127: Implications to Carboxypeptidase A Catalytic ActionsBased on a review of the CPA inhibition, the roles of the hydrophobic pocket of CPA were proposed not only for the recognition of the substrate in the catalysis but also for the release of the product. Discussion of the hydrophobic pocket in the inhibition of the CPA indicated that the induce-fit theory suggested by Koshland takes an important role not only in the recognition of the substrate by CPA but also in the catalytic process.Further establishment of the product inhibition of CPA was taken by designing the amino acid, (2 'S, 2S),3-phenyl-2-(pyrrolidine-2-yl)propionic acid (TM-1). TM-1 was synthesized from L-proline. The appropriate precursor for the subsequent benzylation was obtained using the Ardnt-Eistert reaction as the key step. After transformation to the bicyclo compound 3-6, (4S, 4aS)-4-benzyl-4a, 5, 6, 7-tetrahydro-pyrrolo[1, 2-c]pyrimidine-l, 3-dione, and determination of the absolute configuration of 3-6 using H-H COSY and H-H NOESY technologies, the absolute configuration of TM-1, (2S, 2'S)-3-phenyl-2-(pyrrolidme-2-yI)propionic acid, was correlated. Subsequent kinetic evaluation of TM-1 as CPA inhibitor shown that TM-1 was a very weak inhibitor of CPA through the electro-static interactions between the protonated imino group and the carboxylate of Glu-270. Such resultssuggested that the electro-static interactions should not be considered as the main design rationale for CPA inhibition.Differentiating the chiral circumstance of the active site of CPA was achieved by the different inhibitory activities of all four stereomers of 2-benzyl-3-methanesulfinyl propionic acid, TM-2, which were prepared from diethyl 2-benzylmalonate. The enzymatic resolution of the racemic 2-benzyl-3-methylthiopropionic acid was performed using alpha-chymotrypsin. Following oxidization with sodium periodate gave the diastereomer mixtures, which could be separated by silica gel column chromatography. Hydrolysis was achieved at the neutral condition using Lil in anyhydrous ethyl acetate. Absolute configuration of (25, SR)-TM-2 was assured according to X-ray crystallographies. The stereochemical assignments of the other three isomers of TM-2 were made on the basis of [a] values by comparison to the (25, SR)-TM-2 ([alhpa] = -7.50). Combined with the kinetic results of 2-benzylpentanoic acid and 2-benzyl-3-methylthiopropionic acid, binding of the sulfoxide group of TM-2 to the active site of CPA was confirmed. Accordingly, TM-2 were suggested as the substrate analog inhibitors of CPA, and the sulfoxide group served as an isotererer of a carboxamide moiety of the peptide substrates. Kinetic assayes of the four stereomers of TM-2 shown that the sulfoxide group with S configuration best fit the topologies of the active site of CPA. Thus, the nucleophilic attacking to the substrates was believed from the/e face of the prochiral carbonyl group.Part Two. Arndt-Eistert Reaction of /V-Tosyl Cyclic cr-Amino Acids to a, /3-Unsaturated Esters Bearing a Terminal Tosylamino GroupArndt-Eistert reaction, as one type of Wolff rearrangement, has been widely used in organic synthesis. The Arndt-Eistert reactions of N-tosyl cyclic alpha-amino acids were studied to explore the possible preparation of alpha, beta-unsaturated esters bearing a terminal tosylamino group in one step based on the previously reported preparation of gamma-hydroxy-alpha,beta-unsaturated esters from Arndt-Eistert reaction of alpha, beta-epoxy diazomethyl ketones.L-N-Tosyl aziridine-2-carboxylic acid was prepared from L-serine, and the thermal decomposition of the corresponding alpha-diazomethyl ketone stereospecificly gave the alpha, beta-unsaturated esters with E configuration under the catalysis by silver ion. The concerted mechanism was proposed for the thermally silver ion catalyzed Arndt-Eistert reaction, and the stepwise mechanism of Arndt-Eistert reaction of a, beta-epoxy diazomethyl ketones was also proposed which dif...