Isolation of a pH-Sensitive IgNAR Variable Domain from a Yeast-Displayed, Histidine-Doped Master Library

Marine Biotechnology
Doreen KönningHarald Kolmar

Abstract

In recent years, engineering of pH-sensitivity into antibodies as well as antibody-derived fragments has become more and more attractive for biomedical and biotechnological applications. Herein, we report the isolation of the first pH-sensitive IgNAR variable domain (vNAR), which was isolated from a yeast-displayed, semi-synthetic master library. This strategy enables the direct identification of pH-dependent binders from a histidine-enriched CDR3 library. Displayed vNAR variants contained two histidine substitutions on average at random positions in their 12-residue CDR3 loop. Upon screening of seven rounds against the proof-of-concept target EpCAM (selection for binding at pH 7.4 and decreased binding at pH 6.0), a single clone was obtained that showed specific and pH-dependent binding as characterized by yeast surface display and biolayer interferometry. Potential applications for such pH-dependent vNAR domains include their employment in tailored affinity chromatography, enabling mild elution protocols. Moreover, utilizing a master library for the isolation of pH-sensitive vNAR variants may be a generic strategy to obtain binding entities with prescribed characteristics for applications in biotechnology, diagnostics, and th...Continue Reading

References

Jun 1, 1997·Nature Biotechnology·E T Boder, K D Wittrup
Feb 9, 2000·Biotechnology Progress·J J VanAntwerp, K D Wittrup
Apr 5, 2007·Nature Protocols·Ginger ChaoK Dane Wittrup
Feb 5, 2010·Protein Engineering, Design & Selection : PEDS·Lorenzo BenatuilChung-Ming Hsieh
Jul 19, 2011·Protein Science : a Publication of the Protein Society·Megan L MurtaughJames R Horn
Feb 2, 2012·The Journal of Biological Chemistry·Javier Chaparro-RiggersArvind Rajpal
Jan 2, 2014·Proceedings of the National Academy of Sciences of the United States of America·Eva-Maria StrauchDavid Baker
Jun 26, 2014·Biotechnology Journal·Michael W TraxlmayrChristian Obinger
Aug 16, 2014·Biochimica Et Biophysica Acta·T IgawaK Hattori
Jan 22, 2015·MAbs·Pauline BonvinNicolas Fischer

❮ Previous
Next ❯

Citations

Mar 9, 2017·Protein Science : a Publication of the Protein Society·Thomas Böldicke
Nov 2, 2018·Biochemical Society Transactions·Obinna C UbahCaroline J Barelle
Jan 11, 2018·Microbial Cell Factories·Simon RosowskiSimon Krah
Apr 29, 2020·FASEB Journal : Official Publication of the Federation of American Societies for Experimental Biology·Henk van FaassenGreg Hussack
Jul 14, 2020·Frontiers in Bioengineering and Biotechnology·Steffen C HinzHarald Kolmar
Dec 17, 2020·Frontiers in Immunology·Arturo Macarrón PalaciosHarald Kolmar
Mar 25, 2021·Biological Chemistry·Bernhard ValldorfStefan Zielonka

❮ Previous
Next ❯

Related Concepts

Related Feeds

Bioinformatics in Biomedicine

Bioinformatics in biomedicine incorporates computer science, biology, chemistry, medicine, mathematics and statistics. Discover the latest research on bioinformatics in biomedicine here.