Design of bio-inspired silica-encapsulated protein A for improved immunoprecipitation assays

• Published in: Biochemical engineering journal 2017-12, Vol.128, p.12-18
• Main Authors: Park, Ki Sung, Ki, Mi-Ran, Yeo, Ki Baek, Choi, Jung Hoon, Pack, Seung Pil
• Format: Article
• Language: English
• Subjects: Biosilica; Encapsulation; Immunoprecipitation (IP); Silica-forming peptide (SFP); Staphylococcus aureus Protein A (SpA)
• ISSN: 1369-703X; 1873-295X
• DOI: 10.1016/j.bej.2017.08.017

[Display omitted] •By fusing silica forming peptide (SFP) with Ab-binding Protein A (SpA), SpA-SFPs were newly designed.•SpA-SFPs possessed both of Ab-binding ability and silica forming ability.•SpA-EctP1 has enhanced Ab-binding ability, which is favorable for Ab display.•SpA-SFPs have auto-silicifying ability to construct biosilica-encapsulated SpA particle (SpA-SFP@SiO2).•In IP assay, SpA-SFP@SiO2 showed approximately 300% higher precipitation performance than commercial SpA-particle. Staphylococcus aureus Protein A (SpA) has a high affinity to the Fc region of antibodies (Abs), and SpA-immobilized matrices are widely used for Ab purification or immunoprecipitation (IP) assays. Here, we employed a bio-inspired silica-encapsulation method to improve the Ab-binding ability of an SpA-immobilized matrix. Two silica-forming peptides (SFPs), namely R5 and EctP1, were separately introduced at the C-terminus of SpA to generate two recombinant fusion proteins (SpA-SFPs) with auto-silicifying abilities. When SpA-SFPs were employed as Ab-binders on a solid surface (96-well plate), they showed an effective Ab-binding ability and a better performance than intact SpA. A high binding ability was observed even when an SFP-mediated SpA silica matrix (SpA-SFP@SiO2) was prepared. SpA-SFP@SiO2 showed a higher performance than commercially obtained SpA-Agarose particles and no loss of matrices. Moreover, in IP assays, SpA-SFP@SiO2 showed an approximately 300% higher precipitation of target protein than the commercial SpA-Agarose product when a small amount of cell lysate was used. These findings demonstrated that SpA-SFP could be useful for the development of an efficient immunoassay system.