Protein A-based ligands for affinity chromatography of antibodies

Protein A chromatography is a key technology in the industrial production of antibodies, and a variety of commercial protein A adsorbents are available in shelf. High stability and binding capacity of a protein A adsorbent are two key issues for successful practice of protein A chromatography. Earlier versions of protein A adsorbents ever exhibited serious fragility to typical cleaning-in-place protocols (e.g. washing with sodium hydroxide solution), and suffered from low binding capacity, harsh elution, ligand leakage and other problems involved in industrial applications. During the last three decades, various techniques and approaches have been applied in the improvement of chemical stability and enhancement of binding capacity of protein A-based ligands and adsorbents for antibody purifications. This mini-review focuses on the technical explorations in protein A-based affinity adsorbents, especially protein A-based ligands, including the efforts to increase the chemical stability by site-directed mutations and to improve the binding capacity by ligand polymerization and site-directed immobilization. Moreover, the efforts to develop short peptide ligands based on the structure of protein A, including the biomimetic design strategies and the synthesis of peptide-mixed mode hybrid ligands are discussed. These peptide and peptide-based hybrid ligands exhibit high affinity and selectivity to antibodies, but noteworthy differences in the binding mechanism of antibody from protein A. As a result, bound antibody to the ligands could be effectively eluted under mild conditions. Perspectives for the development of the protein A-based peptide ligands have been extensively discussed, suggesting that the ligands represent a direction for technological development of antibody purification.