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[29] Protein engineering by expressed protein ligation

2000; Academic Press; Linguagem: Inglês

10.1016/s0076-6879(00)28414-0

1557-7988

Ulrich Blaschke, Jonathan Silberstein, Tom W. Muir,

Chemical Synthesis and Analysis

The power of organic chemistry for studying structure–activity relationships in small bioactive peptides is now well established and represents a pillar of medicinal chemistry. Consequently, the application of these same chemical engineering strategies to proteins will have no less an impact in understanding the molecular details of how they work. Going beyond the 20 amino acids dictated by the genetic code permits all manner of modifications to be made in the backbone and amino-acid side chains of a protein, thereby allowing, for example, the precise introduction into proteins of fluorophores, isotopic labels, photoactivable cross-linkers, electron paramagnetic resonance (EPR) spin labels, posttranslational modifications, and countless other unnatural amino acids. Given the possibilities in protein engineering, it is not surprising that an enormous amount of effort has gone into the development of approaches for the site-specific introduction of unnatural amino acids into proteins. Several chemical and biosynthetic strategies are now available for this purpose that, to varying degrees, allow most of the preceding modifications to be incorporated into proteins. This chapter focuses on one such approach, expressed protein ligation (EPL) that allows a target protein to be assembled from a series of recombinant and synthetic polypeptide building blocks.