Journal article
Authors list: Zielonka, S; Weber, N; Becker, S; Doerner, A; Christmann, A; Christmann, C; Uth, C; Fritz, J; Schäfer, E; Steinmann, B; Empting, M; Ockelmann, P; Lierz, M; Kolmar, H
Publication year: 2014
Pages: 236-245
Journal: Journal of Biotechnology
Volume number: 191
ISSN: 0168-1656
DOI Link: https://doi.org/10.1016/j.jbiotec.2014.04.023
Publisher: Elsevier
Abstract:
A novel method for stepwise in vitro affinity maturation of antigen-specific shark vNAR domains is described that exclusively relies on semi-synthetic repertoires derived from non-immunized sharks. Target-specific molecules were selected from a CDR3-randomized bamboo shark (Chiloscyllium plagiosum) vNAR library using yeast surface display as platform technology. Various antigen-binding vNAR domains were easily isolated by screening against several therapeutically relevant antigens, including the epithelial cell adhesion molecule (EpCAM), the Ephrin type-A receptor 2 (EphA2), and the human serine protease HTRA1. Affinity maturation was demonstrated for EpCAM and HTRA1 by diversifying CDR1 of target-enriched populations which allowed for the rapid selection of nanomolar binders. EpCAM-specific vNAR molecules were produced as soluble proteins and more extensively characterized via thermal shift assays and biolayer interferometry. Essentially, we demonstrate that high-affinity binders can be generated in vitro without largely compromising the desirable high thermostability of the vNAR scaffold.
Citation Styles
Harvard Citation style: Zielonka, S., Weber, N., Becker, S., Doerner, A., Christmann, A., Christmann, C., et al. (2014) Shark Attack: High affinity binding proteins derived from shark vNAR domains by stepwise in vitro affinity maturation, Journal of Biotechnology, 191, pp. 236-245. https://doi.org/10.1016/j.jbiotec.2014.04.023
APA Citation style: Zielonka, S., Weber, N., Becker, S., Doerner, A., Christmann, A., Christmann, C., Uth, C., Fritz, J., Schäfer, E., Steinmann, B., Empting, M., Ockelmann, P., Lierz, M., & Kolmar, H. (2014). Shark Attack: High affinity binding proteins derived from shark vNAR domains by stepwise in vitro affinity maturation. Journal of Biotechnology. 191, 236-245. https://doi.org/10.1016/j.jbiotec.2014.04.023
