Review
Engineered affinity proteins—Generation and applications

https://doi.org/10.1016/j.jbiotec.2009.01.014Get rights and content

Abstract

The use of combinatorial protein engineering to design proteins with novel binding specificities and desired properties has evolved into a powerful technology, resulting in the recent advances in protein library selection strategies and the emerge of a variety of new engineered affinity proteins. The need for different protein library selection methods is due to that each target protein pose different challenges in terms of its availability and inherent properties. At present, alternative engineered affinity proteins are starting to complement and even challenge the classical immunoglobulins in different applications in biotechnology and potentially also for in vivo use as imaging agents or as biotherapeutics. This review article covers the generation and use of affinity proteins generated through combinatorial protein engineering. The most commonly used selection techniques for isolation of desired variants from large protein libraries are described. Different antibody derivatives, as well as a variety of the most validated engineered protein scaffolds, are discussed. In addition, we provide an overview of some of the major present and future applications for these engineered affinity proteins in biotechnology and medicine.

Section snippets

Generation of novel affinity proteins by selection from protein libraries

Evolution has generated the great diversity of proteins with all different features required for the processes of life primarily by using the 20 amino acids available in nature. Despite all our efforts to fully understand the nature of proteins, we are still far from being able to create completely new proteins with desired structure and function merely by rational design. By utilizing state-of-the-art in vitro evolution technology it is possible to generate protein variants with new features.

Engineered antibody fragments

Through the evolution the immune system has developed the capability of producing antibodies specific for almost any molecule. Because of their exceptional properties of natural molecular recognition, antibodies have been intensely studied and used in many areas of biological research, biotechnology and for medical diagnostics and therapy. Monoclonal antibodies are excellent affinity proteins, but the intact antibody format suffers from some disadvantages for certain applications. Antibodies

Alternative protein scaffolds

The success of antibody engineering and the increasing experience in the field of combinatorial libraries and protein engineering have inspired researchers to develop new non-immunoglobulin affinity protein lacking the inherent limitations of antibodies. Consequently, today antibodies and antibody derivatives are facing increasing competition from a large number so-called engineered protein scaffolds (Hosse et al., 2006). The term “scaffold” reflects the use of a protein framework that can

Biotechnology research reagents and separation ligands

Engineered alternative scaffolds are in many aspects well suited for biotechnology applications (Fig. 3). Generally, high biophysical stability, long shelf-life and convenient expression at high yields in a prokaryotic host, gives cost-efficient production of engineered affinity proteins, when compared to conventional antibodies. One area of biotechnology applications that have attracted increased attention is the use of engineered affinity proteins as capture agents in affinity matrices. In

Concluding remarks

Engineered affinity proteins are foreseen to have an expanding role in biotechnology, biological science and medical applications in the future. Investigations of independent novel engineered proteins imply that it is possible to generate artificial binding proteins comparable to conventional antibodies in terms of specificity and affinity. However, to truly be able to compete with the well-established antibody technology, engineered protein scaffolds have to display properties superior to that

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