Peptides synthesis racemization, prevention

Carbodiimide peptide synthesis. 1 -Hydroxybenzotriazole (HOBt) is frequently used to suppress racemization in peptide syntheses using carbodiimides. Copper(II) chloride decreases racemization, but also depresses the yield markedly. By judicious use of both additives, racemization can be prevented with only slight effect on the yield. 

Various more or less efficient methods have been reported for the synthesis of 2-(l-ami-noalkyl)thiazole-4-carboxylic acids and their suitably protected derivatives. 237,539,541,558-568 Optimal conditions must be selected in these syntheses to prevent racemization at the chiral aminoalkyl moiety, e.g. when applying a modified Hantzsch synthesis 559 racemization has been observed to occur at the level of the starting Na-protected amino acid thioamide as well as in the base-mediated dehydration step of the intermediate hydroxydihydrothiazoles. 558 The 2-(aminoalkyl)thiazole-4-carboxylic acids are incorporated into the linear precursors by standard procedures of peptide synthesis, 237,514,529,539,552,555,558,564,569 and cyclization is pref-... 

The azide procedure for peptide synthesis and particularly for fragment condensations is considered to be a mainly racemization free method. This low racemization tendency of azides was explained by several theories, which have been reviewed.t l The most plausible cause of racemization is the formation of oxazoles (Scheme 3) and the related enolization. In presence of bases the a-carbon proton is readily abstracted to form an anionic oxazol-5(4//)-one resonance system.For the formation of the oxazol-5(4//)-one the influence of the substituent Y on the a-carbonyl is essential. Since the a-carbonyl group of amino acid azides are less activated and thus relatively insensitive to oxygen containing nucleophiles such as water and alcohols, oxazol-5(4//)-one formation is largely prevented. It was proposed that the soft electron shell of the azide shields the a-carbonyl atom, so that only strong nucleophiles can attack it.t 1 The reactivity towards amines can be explained in a manner analogous to the aminolysis of anchimerically assisted active esters.h 1... 

In peptide synthesis the use of a suitable protection for the N-terminal amino group is required not only to prevent the formation of a complex mixture of oligo- and cyclo-peptides, but an additional demand on the functionality applied for this purpose is that it should prevent possible racemization of the activated amino acid. Racemization usually takes place via an intermediate oxazolone (7) that forms readily from A -acyl-protected amino acids (Scheme 2). This side reaction can be mostly suppressed by using a carbamate as an N-terminal-protecting group. Therefore, nearly all blocking functions currently applied in this field are of the urethane type. 

R ) relative configuration stereosymbol, 360 r synthons. See Radical synthons Racemates, resolution of. See Optical resoiution Racemization prevention in peptide synthesis, 145, 231-232, 234... 

Imidazole protection, temporary protection of the basic and nucleophilic imidazole group of histidine during peptide synthesis. The imidazole moiety has two nonequivalent, but similarly reactive nitrogen atoms, designated tt and r, which cause difficulties in both protection and His racem-ization in peptide synthesis. By application of Boc/Bzl tactics, the jr-benzyloxymethyl (Bom) group is suitable as it prevents racemization and can be cleaved by HE. For Fmoc/Bu chemistry, the best acid-labUe blocking group seems to be r-trityl, as it is stable to bases and cleavable by aqueous TEA at room temperature [T. Brown et al., J. Chem. Soc., Perkin Trans 11982, 1553 P. Sieber, B. Riniker, Tetrahedron Lett. 1987, 28, 6031]. 

At the University of Frankfurt Horst Kessler, now at the Technical University of Munich, can list important achievements in the determination of the 3-dimensional structure of peptides with the help of 2D NMR spectra (cf p. 127). At the nearby Hoechst pharmaceutical company Rolf Geiger (1932-1988) and W. Konig, both former students of F. Weygand, with R. Obermeier and H. Wissmann reported the synthesis of several complex, biologically active peptides, including secretin and insulin. At least equally important are their contributions to the methods of peptide bond formation and protection. The additives proposed by Konig and Geiger for the prevention of racemization (cf p. 87) had a major impact on the practice of peptide synthesis. 

Besides the peptide coupling reactions themselves, also some of the required modification reactions, such as carboxylate group activation and N-terminal amine protection, can also be catalyzed by enzymes. The use of enzymes in peptide synthesis and modification is attractive since it makes side-chain protection superfluous, allows mild reaction conditions that prevent racemization, and can introduce a desired regio-, stereo-, and substrate selectivity. 

In the studies described below cyclic head-to-tail peptides were synthesized by standard solid-phase synthesis utilizing Boc chemistry 27 All syntheses were carried out with Boc-Pro-phenylacetamidomethyl resin. The use of Pro as the C-terminus residue was required so as to prevent racemization during the cyclization reaction (see Section 13.3.1.1). 

Racemization during the synthesis of peptides is a complex problem. The diversity of possible courses followed in the process is compounded by the individuality of the amino acids. This was already shown on the example of S-alkyl-cysteine residues which lose chiral purity by a special mechanism even if protected by a urethane-type protecting group that prevents racemization in other acylamino acids. The opposite end of the scale is represented by proline... 

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