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In-solution antibody harvesting with a plant-produced hydrophobin-Protein A fusion
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A1 Alkuperäisartikkeli tieteellisessä aikakauslehdessä
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en
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11
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Plant Biotechnology Journal, Volume 16, issue 2, pp. 404-414
Abstract
Purification is a bottleneck and a major cost factor in the production of antibodies. We set out to engineer a bifunctional fusion protein from two building blocks, Protein A and a hydrophobin, aiming at low-cost and scalable antibody capturing in solutions. Immunoglobulin-binding Protein A is widely used in affinity-based purification. The hydrophobin fusion tag, on the other hand, has been shown to enable purification by two-phase separation. Protein A was fused to two different hydrophobin tags, HFBI or II, and expressed transiently in Nicotiana benthamiana. The hydrophobins enhanced accumulation up to 35-fold, yielding up to 25% of total soluble protein. Both fused and nonfused Protein A accumulated in protein bodies. Hence, the increased yield could not be attributed to HFB-induced protein body formation. We also demonstrated production of HFBI-Protein A fusion protein in tobacco BY-2 suspension cells in 30 l scale, with a yield of 35 mg/l. Efficient partitioning to the surfactant phase confirmed that the fusion proteins retained the amphipathic properties of the hydrophobin block. The reversible antibody-binding capacity of the Protein A block was similar to the nonfused Protein A. The best-performing fusion protein was tested in capturing antibodies from hybridoma culture supernatant with two-phase separation. The fusion protein was able to carry target antibodies to the surfactant phase and subsequently release them back to the aqueous phase after a change in pH. This report demonstrates the potential of hydrophobin fusion proteins for novel applications, such as harvesting antibodies in solutions.
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Kurppa, K, Reuter, L J, Ritala, A, Linder, M B & Joensuu, J J 2018, 'In-solution antibody harvesting with a plant-produced hydrophobin-Protein A fusion', Plant Biotechnology Journal, vol. 16, no. 2, pp. 404-414. https://doi.org/10.1111/pbi.12780