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The generation of novel recombinant binding proteins will be able to transform cellular and proteomics research, and greatly expand over what is possible with traditional murine monoclonal antibodies. By creating molecules which are fully functional inside the cell, and can be engineered with a variety of functions, they can be used to support such important cell biological activities, and also help structural biology by cocrystallization. Over the last few years, we have developed new classes of synthetic binding proteins that work equally well outside of the cell (like antibodies) as they do inside the cell (unlike antibodies, which largely aggregate in the reducing cytoplasm, as they are dependent on disulfide bonds). Designed Ankyrin Repeat Proteins (DARPins) have been selected to bind to a wide variety of targets with picomolar affinity, and a number of crystal structures of such complexes have been determined. DARPins are very highly expressed, and full consensus proteins are extraordinarily stable. Some examples targeting specific cytoplasmic kinases (distinguishing the phosphorylated and non-phosphorylated state, as well as the kinase), GPCRs and eukaryotic tumor markers will be discussed. More recently, Armadillo Repeat Proteins have been designed for binding to peptides or extended regions from proteins. They hold the promise to develop a modular recognition code, akin to the segmental recognition of one strand of DNA by the other. Modular building blocks might provide the only practical solution to proteome wide protein recognition. The key in all this work has been attention to protein stability in the library design, as all variations introduced for binding specificity will usually destabilize the protein |
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