Protein Expression Analysis

Advances in industry and medicine have led to the engineering of complex “designer” proteins, such as antibodies in targeted therapeutics and enzymes in process development. The ability to easily generate an almost infinite number of variants at the DNA level has increased the demand for improved protein expression methodologies to fully capture what can be produced genetically. Often, the protein of interest is eukaryotic in origin and may require posttranslational modifications specific to its native host or may be toxic to the host cells expressing them. Cell-free protein expression systems have allowed us to step beyond the limits of traditional in vivo expression methodologies by decoupling protein expression from host cell viability (1,2,3). Furthermore, the ability to produce complex proteins using cell-free transcription/translation systems uniquely enables high-throughput directed evolution and protein engineering efforts (4,5). Several cell-free protein expression systems have been developed in the last decade with recent advances focusing on special folding or assembly environments (6,7,8). Equally as important is the capability to transition from the in vitro system to largerscale in vivo expression, while maintaining activity of the target protein (9,10).