Antibody humanization-the Influence of the antibody framework on the CDR-H3 loop ensemble in solution

Protein Engineering, Design & Selection : PEDS
Monica L Fernández-QuinteroKlaus R Liedl

Abstract

Antibody engineering of non-human antibodies has focused on reducing immunogenicity by humanization, being a major limitation in developing monoclonal antibodies. We analyzed four series of antibody binding fragments (Fabs) and a variable fragment (Fv) with structural information in different stages of humanization to investigate the influence of the framework, point mutations and specificity on the complementarity determining region (CDR)-H3 loop dynamics. We also studied a Fv without structural information of the anti-idiotypic antibody Ab2/3H6, because it completely lost its binding affinity upon superhumanization, as an example of a failed humanization. Enhanced sampling techniques in combination with molecular dynamics simulations allow to access micro- to milli-second timescales of the CDR-H3 loop dynamics and reveal kinetic and thermodynamic changes involved in the process of humanization. In most cases, we observe a reduced conformational diversity of the CDR-H3 loop when grafted on a human framework and find a conformational shift of the dominant CDR-H3 loop conformation in solution. A shallow side minimum of the conformational CDR-H3 loop ensemble attached to the murine framework becomes the dominant conformation in s...Continue Reading

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Citations

Jul 29, 2020·Protein Engineering, Design & Selection : PEDS·Monica L Fernández-QuinteroKlaus R Liedl
Aug 1, 2020·Frontiers in Immunology·Monica L Fernández-QuinteroKlaus R Liedl
Aug 28, 2020·Frontiers in Molecular Biosciences·Monica L Fernández-QuinteroKlaus R Liedl
Oct 22, 2020·Communications Biology·Monica L Fernández-QuinteroKlaus R Liedl
Dec 18, 2020·Frontiers in Molecular Biosciences·Monica L Fernández-QuinteroKlaus R Liedl
Jan 22, 2021·Clinical & Translational Oncology : Official Publication of the Federation of Spanish Oncology Societies and of the National Cancer Institute of Mexico·Shuifa WuJieming Xie
Mar 20, 2021·Frontiers in Immunology·Monica L Fernández-QuinteroKlaus R Liedl
May 8, 2021·Frontiers in Molecular Biosciences·Monica L Fernández-QuinteroKlaus R Liedl
Aug 28, 2021·Frontiers in Immunology·Monica L Fernández-QuinteroKlaus R Liedl

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Methods Mentioned

BETA
X-ray

Software Mentioned

cpptraj
PLUMED
pytraj
IgBlast
Protonate3D
PyEMMA
AMBER16
Molecular Operating Environment ( MOE )
Molecular Operating Environment ( MOE
pmemd

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