Enzymatic Synthesis of Peptides

Enzymatic peptide synthesis can proceed by two mechanisms thermodynamic and kinetic control (Kumar and Bhalla 2005). The thermodynamically controlled synthesis of peptides (TCS) with proteases represents the reverse of the hydrolytic breakage of peptide bond catalyzed by those enzymes, as shown in the scheme (Jakubke et al. 1985)  

The kinetically controlled synthesis of peptides (KCS) with proteases can be represented by the following scheme (Bordusa 2002)  

Despite their good catalytic properties, proteases are not ideal catalysts for the synthesis of peptides. Its specificity and selectivity might limit their potential, particularly in the case of rather large peptides where unwanted hydrolytic reactions will occur over the formed product and the substrates. Besides, the use of non-conventional reaction media and the conditions of temperature and pH required for synthesis can be detrimental both for protease activity and stability (Barberis et al. 2002 Bordusa 2002 Quiroga et al. 2005, 2006). However, there are different strategies to overcome such problems, which comprise the engineering of the reaction medium, the biocatalyst and the substrate (Lombard et al. 2005). 

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Peptide Synthesis. The literature on the enzymatic synthesis of peptides is enormous (120—124). Here the basic principles that govern peptide synthesis are illustrated and recent trends in this area reviewed. 

Sauerbrei B,Kappes T,Waldmann H (1997) Enzymatic Synthesis of Peptide Conjugates - Tools for the Study of Biological Signal Transduction. 186 65-86 Sauvage J-P, see Chambron J-C (1993) 165 131-162... 

Enzymatic Synthesis of Peptide Conjugates -Tools for the Study of Biological Signal Transduction... 

Precipitation-driven reactions show some very favorable advantages when compared to other low-water systems for enzymatic synthesis of peptides [55, 56]. 

Schelhaas M, Glomsda S, Hansler M, Jakubke HD, Waldmann H. Enzymatic synthesis of peptides and Ras lipopeptides employing choline ester as a solubilizing, protecting, and activating group. Angew. Chem. Int. Ed. 1996 35 106-109. 

Although, in principal it is possible to perform totally enzymatic synthesis of peptides, in practice combined chemical and enzymatic steps are preferred. For the classical enzymochemical synthesis of polypeptides and even small proteins, the optimum approach is usually synthesis of fragments using the SPPS methodology... 

Bisswanger H (2004) Practical enzymology. Wiley VCH, Weinheim, 272 pp Bjdrup P, Adlercreutz P, Clapes P (1999) Useful methods in enzymatic synthesis of peptides. A comparative study focussing on kinetically controlled synthesis of Ac-Phe-Ala-NHz catalyzed by oc-chymotiipsin. Biocatal Biotransfor 17 319-345 Black M, MiUer R (2006) Platform chemicals from crops. J Chem Technol Biotechnol 81(11) 1725-1728... 

The problem of the enzymatic synthesis of peptide bonds assumed a new aspect in around 1937, when Heinz Fraenkel-Conrat [30], then in the laboratory of M. Bergmann, demonstrated the papain-catalyzed formation at pH 5 of sparingly soluble benzoylglycine anilide from benzoylglycine amide or from benzoylglycine (hippuric acid) and aniline as well as the condensation of benzoylleucine with leucine anilide yielding the nearly insoluble dipeptide benzoyldileucine anilide (Fig. 6). 

M. Bergmann, H. Fraenkel-Conrat, The enzymatic synthesis of peptide bonds, J. Biol. Chem. 124 1-6 (1938)... 

The enzymatic synthesis of peptides (Scheme 6.24) from which proteins can be constructed is not so limited, and chemical synthesis has an even wider application, but these are not yet suitable techniques for manufacture. Moreover, there are no general methods for building the peptides into full protein structures. Nevertheless, enzymes do have a role in the manufacture of peptides themselves. In a mixture of butan-l,4-diol and water, trypsin will catalyse the exchange of the carboxy-terminal alanine of porcine insulin with threonine t-butyl ester (Scheme 6.25). The reaction is essentially a transpeptidation in which the acyl group of lysine is transferred from one amino group on alanine to another on the threonine. This converts porcine insulin into the ester of the human hormone, and a simple deprotection yields one of the commercial products. 

A new type of enzymatic synthesis of peptides has been found by Brenner et Esters of DL-methionine or DL-threonine incubated... 

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