Racemization Mechanism of Cysteine Dipeptide Active Ester Derivatives

Article abstract of DOI:10.1021/jo00146a032

The racemization rate constants of Z-Gly-L-Cys(Bzl)-OH and Z-L-Cys(Bzl)-OH active esters were practically identical, even though glycyl dipeptide active esters usually racemize 50-100 times faster than the amino acid active esters.This observation led to the conclusion that Z-Gly-L-Cys(Bzl)-ONp and other active estrs do not racemize through the 5(4H)-oxazolone but through an enolization mechanism.The deuterium isotope effect was used to demonstrate that the removal of the α-hydrogen in the racemization is indeed the rate-determining step.Ac-Cys(Bzl)-OH was convertedwith DCC to the 5(4H)-oxazolone and deuterated with AcO²H: during this procedure Ac2O and Ac-(α-²H)-DL-Cys(Bzl)-OH were formed, the latter was resolved with acylase I in the presence of Co(2+) ions.H(α-²H)-L-Cys(Bzl)-OH (II) was converted through Z-(α-²)-L-Cys(Bzl)-OH (V), Z-(α-²H)-L-Cys(Bzl)-ONp (VI), and HBr*H-(α-²H)-L-Cys(Bzl)-ONp (VII) to Z-Gly-(α-²H)-L-Cys(Bzl)-ONp (VIII).The racemization rate constant k_r^2H for deuterated VIII in THF with Et3N was 335 * 10^-6 M^-1 s^-1, while the k_τ(H) for the undeuterated VIII was 704 * 10^-6 M^-1 s^-1, indicating an isotope effect of 2.1.The 5(4H)- and 5(4²H)-oxazolones, XII and XIII, were prepared from Z-Gly-L-Cys(Bzl)-OH and Z-Gly-(α-²H)-L-Cys(Bzl)-OH with DCC under controlled conditions in 73percent optical purity.Racemization of XII and XIII with Et3N in THF is instantaneous while the coupling rate with H-Val-OMe is k_c = 8.5*10^-2 M^-1 s^-1; that is, k_r >> k_c.The racemization rate of 5(4H)-oxazolone XII during coupling with H-Val-OMe is 0.34 M^-1 s^-1, while the autoracemization rate of XII is 0.08 M^-1 s^1-.These results support the conclusions that Z-Gly-Cys(Bzl)-ONp racemizes mainly through an enolization mechanism and that the cysteine side chain is responsible for this abnormal behavior.It is expected in practical peptide synthesis that coupling cysteine dipeptide active esters will yield an optically purer product than the coupling of cysteine active ester derivatives.