Antigen-binding affinity and thermostability of chimeric mouse-chicken IgY and mouse-human IgG antibodies with identical variable domains

Juho Choi,Jin-Kyoo Kim,Yeonkyoung Ham,Jihyun Lee,Myung-Hee Kwon,Jeonghyun Lee,Joungmin Lee,Minjae Kim,Youngsil Seo

doi:10.1038/s41598-019-55805-4

Juho Choi, Jin-Kyoo Kim + Show 7 more

Open Access

https://doi.org/10.1038/s41598-019-55805-4

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Journal: Scientific Reports	Publication Date: Dec 1, 2019
Citations: 7	License type: open-access

Affiliation: Ajou University, Changwon National University

Abstract

Constant (C)-region switching of heavy (H) and/or light (L) chains in antibodies (Abs) can affect their affinity and specificity, as demonstrated using mouse, human, and chimeric mouse-human (MH) Abs. However, the consequences of C-region switching between evolutionarily distinct mammalian and avian Abs remain unknown. To explore C-region switching in mouse-chicken (MC) Abs, we investigated antigen-binding parameters and thermal stability of chimeric MC-6C407 and MC-3D8 IgY Abs compared with parental mouse IgGs and chimeric MH Abs (MH-6C407 IgG and MH-3D8 IgG) bearing identical corresponding variable (V) regions. The two MC-IgYs exhibited differences in antigen-binding parameters and thermal stability from their parental mouse Abs. However, changes were similar to or less than those between chimeric MH Abs and their parental mouse Abs. The results demonstrate that mammalian and avian Abs share compatible V-C region interfaces, which may be conducive for the design and utilization of mammalian-avian chimeric Abs.

Highlights

Antibodies (Abs) are composed of two identical heavy (H) chains and two identical light (L) chains
The two chimeric MH-IgGs were purified from the culture supernatant of FreeStyle 293-F cells transfected with the specific KV10 vectors using Protein A-agarose resin at 7 days post-transfection, and the two chimeric MC-IgYs were purified using Protein L-agarose
SDS-PAGE analysis showed that mouse IgGs (6C407 and 3D8) and MH-IgGs (6C407 and 3D8) were of the expected sizes (~50 and ~25 kDa, respectively) under reducing conditions and >150 kDa under non-reducing conditions (Fig. 2a)

Summary

Introduction

Antibodies (Abs) are composed of two identical heavy (H) chains and two identical light (L) chains. During C → V allosteric signaling, C-domain switching of Abs frequently affects the conformation of the variable (V) region, leading to differences in antigen-binding parameters of V domains, thermodynamics, and functional efficacy[2,3]. This indicates the possibility for affinity modulation through isotype switching without engineering of V domains. The consequences of C-domain switching between mammalian and non-mammalian Abs, including avian Abs, remain unknown Both humans and mice express Abs with five classes of H chain (μ, δ, γ, ε, and α) and two classes of L chain (κ and λ) comprising different IgM, IgD, IgG, IgE, and IgA isotypes.

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