Expanding the Genetic Code in Vitro and in Vivo.

Insight into biological function at almost every level, from catalysis to signal transduction to structure, requires a detailed understanding of proteins, biopolymers of remarkable di versity assembled from only twenty amino acid building blocks. Site-directed mutagenesis - the process by which an amino acid in a protein is swapped for one of the other 19 natural proteinogenic amino acids - has emerged as one of the most useful and powerful tools at the biochemists disposal. Not only does site-directed mutagenesis allow the identification of residues critical for catalysis, binding, folding, or structure, but it also made possible the first protein engineering experiments. The bioorganic chemist, however, cannot be fully satisfied with this method because the changes allowed are very limited compared to the physical or-ganic chemists ability to subdy alter steric or electronic properties, or the synthetic chemist's ability to install useful functionalities or unique chemical handles for elaboration. Methods to alter proteins in more general ways have been developed over the last decade, inspiring the notion of "unnatural" life forms-living cells capable of using amino acids not accessible to life as we have observed it. Todays powerful in vitro methods of unnatural protein mutagenesis have become increasingly useful and accessible and have enhanced our understanding of pro-tein fiinction. In addition, the advent of living cells producing proteins with unnatural amino acids will allow a level of biophysical and cell-biological characterization that would have been difficult to imagine a decade ago. [ABSTRACT FROM AUTHOR]