Towards engineering proteins by site-directed incorporation in vivo of non-natural amino acids.

Altering protein structure via the techniques of protein engineering has already allowed the development of proteins displaying both modified and novel activities. The only limitation of conventional site-directed mutagenesis, the cornerstone of protein engineering, is that substitutions are restricted to the 20 naturally occurring, proteinogenic amino acids. However, the discovery of a 21st amino acid, selenocysteine, and the development of novel in vitro translation systems have demonstrated that considerably more substitutions are possible. To this end, a number of experimental approaches have been developed that allow the incorporation of synthetic amino acids into proteins. Some of these have already been successfully applied in vitro and efforts to transfer this technology to in vivo systems are now underway.