title: Engineering a high-affinity anti-IL-15 antibody: crystal structure reveals an α-helix in VH CDR3 as key component of paratope.
abstract: Interleukin (IL) 15 is an inflammatory cytokine that plays an essential role in the activation, proliferation, and maintenance of specific natural killer cell and T-cell populations, and has been implicated as a mediator of inflammatory diseases. An anti-IL-15 antibody that blocked IL-15-dependent cellular responses was isolated by phage display and optimised via mutagenesis of the third complementarity-determining regions (CDRs) of variable heavy (VH) and variable light chains. Entire repertoires of improved variants were recombined with each other to explore the maximum potential sequence space. DISC0280, the most potent antibody isolated using this comprehensive strategy, exhibits a 228-fold increase in affinity and a striking 40,000-fold increase in cellular potency compared to its parent. Such a wholesale recombination strategy therefore represents a useful method for exploiting synergistic potency gains as part of future antibody engineering efforts. The crystal structure of DISC0280 Fab (fragment antigen binding), in complex with human IL-15, was determined in order to map the structural epitope and paratope. The most remarkable feature revealed lies within the paratope and is a novel six-amino-acid α-helix that sits within the VH CDR3 loop at the center of the antigen binding site. This is the first report to describe an α-helix as a principal component of a naturally derived VH CDR3 following affinity maturation.
authors.affiliation: MedImmune Ltd., Milstein Building, Granta Park, Cambridge CB1 6GH, UK. lowed@medimmune.com
authors.forename: David C
authors.initials: DC
authors.lastname: Lowe
authors.forename: Stefan
authors.initials: S
authors.lastname: Gerhardt
authors.forename: Alison
authors.initials: A
authors.lastname: Ward
authors.forename: David
authors.initials: D
authors.lastname: Hargreaves
authors.forename: Malcolm
authors.initials: M
authors.lastname: Anderson
authors.forename: Franco
authors.initials: F
authors.lastname: Ferraro
authors.forename: Richard A
authors.initials: RA
authors.lastname: Pauptit
authors.forename: Debbie V
authors.initials: DV
authors.lastname: Pattison
authors.forename: Catriona
authors.initials: C
authors.lastname: Buchanan
authors.forename: Bojana
authors.initials: B
authors.lastname: Popovic
authors.forename: Donna K
authors.initials: DK
authors.lastname: Finch
authors.forename: Trevor
authors.initials: T
authors.lastname: Wilkinson
authors.forename: Matthew
authors.initials: M
authors.lastname: Sleeman
authors.forename: Tristan J
authors.initials: TJ
authors.lastname: Vaughan
authors.forename: Philip R
authors.initials: PR
authors.lastname: Mallinder
mesh_terms: D000595:Amino Acid Sequence; D057134:Antibodies, Neutralizing / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification*; D001666:Binding Sites, Antibody / Q000235:genetics; D022801:Complementarity Determining Regions / Q000737:chemistry* / Q000235:genetics; D000939:Epitopes / Q000737:chemistry / Q000235:genetics; D006801:Humans; D007140:Immunoglobulin Fab Fragments / Q000737:chemistry / Q000235:genetics / Q000276:immunology; D007143:Immunoglobulin Heavy Chains / Q000737:chemistry / Q000235:genetics / Q000276:immunology; D007147:Immunoglobulin Light Chains / Q000737:chemistry / Q000235:genetics / Q000276:immunology; D019409:Interleukin-15 / Q000037:antagonists & inhibitors* / Q000276:immunology*; D008969:Molecular Sequence Data; D009154:Mutation; D015202:Protein Engineering
chemical_list: D057134:Antibodies, Neutralizing; D022801:Complementarity Determining Regions; D000939:Epitopes; D007140:Immunoglobulin Fab Fragments; D007143:Immunoglobulin Heavy Chains; D007147:Immunoglobulin Light Chains; D019409:Interleukin-15