title: Mutational landscape of antibody variable domains reveals a switch modulating the interdomain conformational dynamics and antigen binding.
abstract: Somatic mutations within the antibody variable domains are critical to the immense capacity of the immune repertoire. Here, via a deep mutational scan, we dissect how mutations at all positions of the variable domains of a high-affinity anti-VEGF antibody G6.31 impact its antigen-binding function. The resulting mutational landscape demonstrates that large portions of antibody variable domain positions are open to mutation, and that beneficial mutations can be found throughout the variable domains. We determine the role of one antigen-distal light chain position 83, demonstrating that mutation at this site optimizes both antigen affinity and thermostability by modulating the interdomain conformational dynamics of the antigen-binding fragment. Furthermore, by analyzing a large number of human antibody sequences and structures, we demonstrate that somatic mutations occur frequently at position 83, with corresponding domain conformations observed for G6.31. Therefore, the modulation of interdomain dynamics represents an important mechanism during antibody maturation in vivo.
authors.affiliation: Department of Antibody Engineering, Genentech Research and Early Development, Genentech, Inc., South San Francisco, CA 94080; koenig.patrick@gmail.com germainefuh@gmail.com.
authors.forename: Patrick
authors.initials: P
authors.lastname: Koenig
authors.affiliation: Department of Antibody Engineering, Genentech Research and Early Development, Genentech, Inc., South San Francisco, CA 94080.
authors.forename: Chingwei V
authors.initials: CV
authors.lastname: Lee
authors.affiliation: Department of Early-Stage Pharmaceutical Development, Pharma Technical Development, Genentech, Inc., South San Francisco, CA 94080.
authors.forename: Benjamin T
authors.initials: BT
authors.lastname: Walters
authors.affiliation: Department of Molecular Biology, Genentech Research and Early Development, Genentech, Inc., South San Francisco, CA 94080.
authors.forename: Vasantharajan
authors.initials: V
authors.lastname: Janakiraman
authors.affiliation: Department of Molecular Biology, Genentech Research and Early Development, Genentech, Inc., South San Francisco, CA 94080.
authors.forename: Jeremy
authors.initials: J
authors.lastname: Stinson
authors.affiliation: Department of Early-Stage Pharmaceutical Development, Pharma Technical Development, Genentech, Inc., South San Francisco, CA 94080.
authors.forename: Thomas W
authors.initials: TW
authors.lastname: Patapoff
authors.affiliation: Department of Antibody Engineering, Genentech Research and Early Development, Genentech, Inc., South San Francisco, CA 94080; koenig.patrick@gmail.com germainefuh@gmail.com.
authors.forename: Germaine
authors.initials: G
authors.lastname: Fuh
mesh_terms: D000906:Antibodies / Q000737:chemistry / Q000235:genetics* / Q000276:immunology; D000915:Antibody Affinity; D000941:Antigens / Q000276:immunology; D001666:Binding Sites, Antibody; D006801:Humans; D007140:Immunoglobulin Fab Fragments / Q000737:chemistry / Q000235:genetics* / Q000276:immunology; D009154:Mutation; D011487:Protein Conformation
chemical_list: D000906:Antibodies; D000941:Antigens; D007140:Immunoglobulin Fab Fragments
keywords: affinity maturation; antibody; conformational dynamics; deep mutational scanning; immunology