Enlightening the Fcab-antigen interaction via X-ray crystallography & solution studies

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00:00:00 Welcome
00:00:22 Elisabeth Lobner
00:00:33 Enlightening the Fcab-antigen interaction via X-ray crystallography & solution studies
00:01:41 Enlightening the Fcab-antigen interaction via X-ray crystallography & solution studies
00:02:16 Projects
00:02:54 Introduction: Modular Antibody Technology – How to get an Fcab?
00:03:01 IgG1-Fc binds them all
00:03:50 Modular Antibody Engineering
00:04:15 Modular Antibody Engineering
00:04:48 Fcabs under study
00:06:21 Fcabs under study
00:07:02 Project I: Fcab-HER2 interaction: A ménage à trois
00:07:02 X-ray structures of Fcabs
00:07:16 X-ray structures of Fcabs
00:08:09 X-ray structures of Fcabs
00:10:04 HER2
00:11:48 Targeting HER2
00:12:34 SEC-MALS
00:13:10 SEC-MALS
00:13:23 SEC-MALS
00:13:59 X-ray structure of Fcab‒HER2 complexes
00:14:45 X-ray structure of H10-03-6‒HER2 complex
00:15:20 ITC measurement of STAB19-HER2 interaction
00:15:43 Discrepancy in binding stoichiometries
00:15:50 Discrepancy in binding stoichiometries
00:16:37 FCS measurements of STAB19-HER2 interaction
00:17:50 Negative cooperative binding behaviour
00:19:05 Project II: Fcab-VEGF interaction: Generation of a two-faced Fcab
00:19:17 X-ray structure of CT6
00:19:49 X-ray structure of CT6
00:20:41 VEGF
00:21:46 Targeting VEGF
00:22:18 SEC-MALS
00:22:33 SEC-MALS
00:23:22 SEC-MALS
00:23:50 Generation of heterodimeric CT6
00:24:49 Generation of heterodimeric CT6
00:24:54 SEC-MALS
00:25:09 SEC-MALS
00:25:34 X-ray structure of the JanusCT6-VEGF complex
00:26:05 ITC measurement of JanusCT6-VEGF interaction
00:27:08 Summary & Conclusion
00:30:25 Summary & Conclusion
00:31:11 Thank You...
00:31:19 Thank You...
00:31:27 Question & Answer

The modular nature of antibodies allows the design of antibody-related formats with tailored characteristics. The antibody’s Fc part, encompassing the effector functions, represents an attractive scaffold for engineering therapeutic molecules as demonstrated by the generation of Fcabs (Fc domain with antigen-binding sites). By engineering the C-terminal loops (AB, CD and EF loop) in the CH3 domains, two antigen binding sites can be inserted in close proximity.

The study presented in this webinar deals with the determination of the first X-ray structures of three Fcabs that differ in loop design, specificity, affinity and thermal stability and reveals their overall structural integrity and native fold.

Furthermore, by investigation of the interaction of these Fcabs with their respective antigens (HER2 or VEGF) using a set of complementary techniques (X-ray crystallography, size exclusion chromatography combined with multi-angle light scattering [SEC-MALS], isothermal titration calorimetry [ITC], fluorescence correlation spectroscopy [FCS]), insights into the binding modes and binding stoichiometries were gained.


Elisabeth Lobner is a post-doctoral researcher in the Department of Biotechnology at the University of Natural Resources and Life Sciences (BOKU) in Vienna, Austria. Her current research mainly focuses on the application of animal cell factories, particularly on difficult-to-express proteins. During her 4-year tenure as a PhD student at the Department of Chemistry (BOKU) in cooperation with F-star Biotechnology Ltd., she investigated the interaction between certain antibody-fragments and their respective antigens. Before this, Elisabeth completed her Master’s degree in biotechnology at BOKU.


Who should attend?
Everyone who focuses on antibodies and antibody fragments and is interested in state-of-the-art biophysical methods to analyze protein-protein interactions.

What will you learn?
The presented study demonstrates how to apply and exploit complementary biophysical methods to shed light on protein-protein interactions.