Peptides synthesis, racemization

Carbamates can be used as protective groups for amino acids to minimize racem-ization in peptide synthesis. Racemization occurs during the base-catalyzed coupling reaction of an A-protected, carboxyl-activated amino acid and takes place in the intermediate oxazolone that forms readily from an A-acyl-protected amino acid (R = alkyl, aryl) ... 

Carbodiimide peptide synthesis. Racemization often presents a problem in the carbodiimide method (1, 233). Several additives have been recommended, such as N-hydroxysuccinimide and 1-hydroxybenzotriazole (3, 156 5, 342). Some Levris acids have also been found to suppress racemization SbCls is very effective, but yields of peptides are low. The most useful additive appears to be ZnClj, and this salt also increases the yield. ... 

With the dicyclohexylcarbodiimide (DCQ reagent racemization is more pronounced in polar solvents such as DMF than in CHjCl2, for example. An efficient method for reduction of racemization in coupling with DCC is to use additives such as N-hydroxysuccinimide or l-hydroxybenzotriazole. A possible explanation for this effect of nucleophilic additives is that they compete with the amino component for the acyl group to form active esters, which in turn reaa without racemization. There are some other condensation agents (e.g. 2-ethyl-7-hydroxybenz[d]isoxazolium and l-ethoxycarbonyl-2-ethoxy-l,2-dihydroquinoline) that have been found not to lead to significant racemization. They have, however, not been widely tested in peptide synthesis. 

The major disadvantage of solid-phase peptide synthesis is the fact that ail the by-products attached to the resin can only be removed at the final stages of synthesis. Another problem is the relatively low local concentration of peptide which can be obtained on the polymer, and this limits the turnover of all other educts. Preparation of large quantities (> 1 g) is therefore difficult. Thirdly, the racemization-safe methods for acid activation, e.g. with azides, are too mild (= slow) for solid-phase synthesis. For these reasons the convenient Menifield procedures are quite generally used for syntheses of small peptides, whereas for larger polypeptides many research groups adhere to classic solution methods and purification after each condensation step (F.M. Finn, 1976). 

Diethylphosphoryl cyanide 3 as a reagent lor amide bond lormation and applicallon to peptide synthesis tree ol racemization. 

OS 25] [R 4] [P 17] For dipeptide formation from the pentafluorophenyl ester of (J )-2-phenylbutyric acid and (S)-a-methylbenzylamine an extent of racemization of 4.2% was found [86]. At higher concentration (0.5 instead of 0.1 M), a higher degree of racemization was found (7.8%). This experiment also served to demonstrate monitoring of the racemization of a simple carboxylic acid used in peptide synthesis. 

Watts, P., Wiles, C., Haswell, S. )., PoMBO-ViLLAR, E., Investigation of racemization in peptide synthesis within a micro reactor. Lab. Chip 2 (2002) 141-144. 

A second strategy is to attach a linker (also referred to as a handle or anchor) to the resin followed by assembly of the molecule. A linker is bifunctional spacer that serves to link the initial synthetic unit to the support in two discrete steps (Fig. 3). To attach a linker to a chloromethyl-PS resin, a phenol functionality such as handle 4 is used to form an ether bond (Fig. 4). To attach the same handle to an amino-functionalized support, acetoxy function 5 or a longer methylene spacer of the corresponding phenol is applied to form an amide bond. Both of these resins perform similarly and only differ in their initial starting resin [4], An alternative approach is to prepare a preformed handle in which the first building block is prederivatized to the linker and this moiety is attached to the resin. For peptide synthesis, this practice is common for the preparation of C-terminal peptide acids in order to reduce the amount of racemization of the a-carbon at the anchoring position [5],... 

Bernatowicz MS, Kearney T, Neves RS, Koster H. An efficient method for racemization-free attachment of 9-fluorenylmethyloxycarbonyl-amino acids to peptide synthesis supports. Tetrahedron Lett 1989 30 4341 4344. 

Application of amino acid silyl esters or A-silyl amino acid silyl esters as amino components is very convenient in peptide synthesis with CDI, because the resulting peptide silyl esters are easily hydrolyzed to dipeptides during the usual work up. They need not be saponified in a separate step, as would be the case with the corresponding alkyl esters. Furthermore, no racemization occurs with this method.tl8],tl9]... 

Another -activation of amino acids for peptide synthesis is achieved by preparing sulfenamides from sulfenylimidazoles. A sulfenylimidazole is formed in situ from the sulfenyl chloride (prepared from the disulfide and chlorine) and imidazole, which reacts further with an amino acid ester to give a sulfenamide in high yield. Conversion of such sulfenamides with IV-acyl amino acids by means of triphenylphosphine affords dipeptides with racemization of less than 0.5%.[481... 

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. 

M Williams, GT Young. Further studies on racemization in peptide synthesis, in GT Young, ed. Peptides 1962. Proceedings of the 5th European Peptide Symposium, Pergamon, Oxford, 1963, pp 119-121. 

NL Benoiton. Quantitation and the sequence dependence of racemization in peptide synthesis, in E Gross, J Meienhofer, eds. The Peptides Analysis, Synthesis, Biology, Academic, New York, 1981, Vol 5, pp 341-361. 

FMF Chen, Y Lee, R Steinauer, NL Benoiton. Mixed anhydrides in peptide synthesis. Reduction of urethane formation and racemization using A-methylpiperidine as tertiary amine base. J Org Chem 48, 2939, 1983. 

JR McDermott, NL Benoiton. JV-Methylamino acids in peptide synthesis. III. Racemization during deprotection by saponification and acidolysis. Can J Chem 51, 2555,... 

B Liberek. The nitrile group in peptide chemistry. V. Racemization during peptide synthesis. 4. Racemization of active esters of phthaloyl-P-cyano-L-alanine in the presence of trie thy lamine. Acad Pol Sci Ser Sci Chim 11, 677, 1963. 

M Goodman, L Levine. Peptide synthesis via active esters. IV. Racemization and ring-opening reactions of optically active oxazolones. JAm Chem Soc 86, 2918, 1964. 

F Weygand, A Prox, L Schmidhammer, W Konig. Gas chromatographic investigation of racemization in peptide synthesis. Angew Chem 75, 282, 1963. 

M Bodanszky, LE Conklin. A simple method for the study of racemization in peptide synthesis. Chem Commun 773, 1967. 

N Izumiya, M Muraoka. Racemization test in peptide synthesis. J Am Chem Soc 91, 2391, 1969. 

B Weinstein, AE Pritchard. Amino-acids and Peptides. Part XXVIII. Determination of racemization in peptide synthesis by nuclear magnetic resonance spectroscopy. J Chem Soc Perkin Trans 1, 1015, 1972. 

NL Benoiton, K Kuroda, FMF Chen. Racemization in peptide synthesis. A laboratory experiment for senior undergraduates. Int J Pept Prot Res 15, 475, 1980. 

JG Adamson, T Hoang, A Crivici, GA Lajoie. Use of Marfey s reagent to quantitate racemization upon anchoring of amino acids to solid supports for peptide synthesis. Anal Biochem 202, 210, 1992. 

SJ Harding, I Heslop, JH Jones, ME Wood. The racemization of histidine in peptide synthesis. Further studies, in HLS Maia, ed. Peptides 1994. Proceedings of the 23rd European Peptide Symposium, Escom, Leiden, 1995, pp 189-190. 

T Miyazawa, T Otomatsu, Y Fukui, T Yamada, S Kuwata. Racemization-free and efficient peptide synthesis by the carbodiimide method using 1-hydroxybenzotriazole and copper(II) chloride simultaneously as additives. J Chem Soc Chem Commun 419, 1988. 

T Miyazawa, T Donkai, T Yamada, S Kuwata. Effect of copper(II) chloride on suppression of racemization in peptide synthesis by the mixed-anhydride and related methods. Int J Pept Prot Res 40, 49, 1992. 

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