Chemical synthesis of proteins

Biological synthesis of proteins is a complex process requiring ordered macromolecular surfaces, protein catalysts, energy storage forms, etc. This alone would indicate that the chemical synthesis of a protein would be a difficult undertaking. Therefore, a number of considerations must be made before attempting such a project. 

specificity or sequential coupling of the amino acids must be achieved. Unwanted side reactions must be avoided. Consider, for example, the synthesis of the dipeptide glycylalanine (Gly-Ala). It is not possible to mix the two amino acids together and let them react as the desired sequence will not be the only product. For example, it is possible that glycine will 

Another factor peculiar to biological systems must be considered optical purity. Proteins are made of L-amino acids. As such, a chemical synthesis must start with L-amino acids and racemization must be minimized during the synthesis. This is especially true in the synthesis of an enzyme, as catalytic activity is dependent upon optical integrity. Amino acids are particularly susceptible to racemization once they have been acylated (i.e., addition of an acyl protecting group to the amino function) via intermediate azlactone 

The a-proton of the azlactone is quite base labile (stabilized carbanion) so that a proton may be lost, but can later add to either side of the azlactone plane with subsequent loss of optical purity. Further, the azlactone may participate in peptide bond forming reactions, so that it disappears during the synthesis, but racemic amino acids are introduced into the product protein. 

This has been replaced by the less reactive tert-BOC azide. The azide has been used extensively until recently when it was reported to be dangerous (explosive). Presently, the reagents of choice are BOC-ON [2-(tert-but-oxycarbonyloxyimino)-2-phenylacetonitrile] and di-tert-butyl dicarbonate. Both give tert-BOC amino acids in high yields. 

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Nilsson BL, Soellner MB, Raines RT (2005) Chemical synthesis of proteins. Annu Rev Biophys Biomol Struct 34 91-118... 

This strategy was first realized by Lozinsky et al., who studied the redox-initiated free-radical copolymerization of thermosensitive N-vinylcaprolactam with hydrophilic N-vinylimidazole at different temperatures, as well as by Chi Wu and coworkers. Lozinsky presents an extensive review of the experimental approaches, both already described in the literature and potential new ones, to chemical synthesis of protein-like copolymers capable of forming core-shell nanostructures in a solution. 

Approaches to Chemical Synthesis of Protein-Like Copolymers... 

Abstract Protein-like copolymers were first predicted by computer-aided biomimetic design. These copolymers consist of comonomer units of differing hydrophilicity/hydro-phobicity. Heterogeneous blockiness, inherent in such copolymers, promotes chain folding with the formation of specific spatial packing a dense core consisting of hydrophobic units and a polar shell formed by hydrophilic units. This review discusses the approaches, those that have already been described and potential approaches to the chemical synthesis of protein-like copolymers. These approaches are based on the use of macromolecular precursors as well as the appropriate monomers. In addition, some specific physicochemical properties of protein like copolymers, especially their solution behaviour in aqueous media, are considered. 

On the basis of general considerations, it is quite evident that the chemical synthesis of protein-like HP-copolymers can be performed by using one of the following three different methods ... 

Lozinsky V.I. Approaches to Chemical Synthesis of Protein-Like Copolymers. Vol. 196, pp. 87-128. 

S Sakakibara. Chemical synthesis of proteins in solution. Biopolymers (Pept Sci) 51, 279, 1999. 

H Yajima, N Fujii, S Funakoshi, T Watanabe, E Murayama, A Otaka. New strategy for the chemical synthesis of proteins. Tetrahedron, 44, 805-819, 1988. 

Semisynthesis can be defined as the use of fragments of proteins, or intact proteins themselves, as ready-made intermediates in the chemical synthesis of proteins. Many modifications to the native structure of a protein can be expected to have profound effects, and produce materials of the greatest academic or practical value, but only involve a change of one or a few atoms out of the thousands that often go to make up the complete molecule. If a semisynthetic method can be found to incorporate, at the correct site, a small synthetic fragment carrying the wanted change, there is no need to go to the trouble of constructing the rest of the molecule when it has already been preformed for us biosynthetically. 

Second, researchers can synthesize proteins without the use of living systems at all, employing only chemical techniques familiar to any chemist. A number of techniques have been developed for the chemical synthesis of proteins. Perhaps the simplest and most obvious (but definitely not the fastest) has been called stepwise synthesis. In this procedure, amino acids are added to each other, one at a... 

Previously, Wlodawer and coworkers (1989) had reported the complete chemical synthesis of HIV protease from L-amino acids (the L-enzyme), using the process shown in Figure 3-29. Normal HIV protease contains two Cys residues at positions 67 and 95. Because chemical synthesis of proteins containing Cys is technically difficult, Wlodawer and colleagues substituted the synthetic amino acid L-a-amino-TC-butyric acid (Aba) for the two Cys residues in the protein. In the authors words, this was done so as to reduce synthetic difficulties associated with Cys deprotection and ease product handling. ... 

Solid-phase synthesis seemed to be the key to a fascinating possibility the chemical synthesis of proteins able to function. Maybe this El Dorado ... 

Nilsson BL, Soellner MB, Raines RT. Chemical synthesis of proteins. Annu. Rev. Biophys. Biomol. Struct. 2005 34 91-118. Pegoraro L, Moroder L. Synthesis of lipopeptides. In Houben-Weyl Methods of Organic Chemistry, Synthesis of Peptides and Peptidomimetics. Goodman M, Felix A, Moroder L, Toniolo C, eds. 2003. Thieme, Stuttgart, Germany. 

Fragment condensation on the solid support has been shown to be a feasible approach towards chemical synthesis of proteins. A progress report on the example of the TCR p-chain and AFP type III has been given. In the latter case a pure AFP type III peptide was obtained that however did not fold properly. 

J.A. Borgia and GB. Fields. 2000. Chemical synthesis of proteins Trends Biotechnol. 18 243-251. (PubMed)... 

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