De novo design of antibody complementarity determining regions binding a FLAG tetra-peptide

Kevin C. Entzminger, Jeong Min Hyun, Robert J. Pantazes, Athena C. Patterson-Orazem, Ahlam N. Qerqez, Zach P. Frye, Randall A. Hughes, Andrew D. Ellington, Raquel L. Lieberman, Costas D. Maranas, Jennifer A. Maynard

Research output: Contribution to journalArticle

7 Citations (Scopus)

Abstract

Computational antibody engineering efforts to date have focused on improving binding affinities or biophysical characteristics. De novo design of antibodies binding specific epitopes could greatly accelerate discovery of therapeutics as compared to conventional immunization or synthetic library selection strategies. Here, we employed de novo complementarity determining region (CDR) design to engineer targeted antibody-antigen interactions using previously described in silico methods. CDRs predicted to bind the minimal FLAG peptide (ASP-Tyr-Lys-ASP) were grafted onto a single-chain variable fragment (scFv) acceptor framework. Fifty scFvs comprised of designed heavy and light or just heavy chain CDRs were synthesized and screened for peptide binding by phage ELISA. Roughly half of the designs resulted in detectable scFv expression. Four antibodies, designed entirely in silico, bound the minimal FLAG sequence with high specificity and sensitivity. When reformatted as soluble antigen-binding fragments (Fab), these clones expressed well, were predominantly monomeric and retained peptide specificity. In both formats, the antibodies bind the peptide only when present at the amino-terminus of a carrier protein and even conservative peptide amino acid substitutions resulted in a complete loss of binding. These results support in silico CDR design of antibody specificity as an emerging antibody engineering strategy.

Original languageEnglish (US)
Article number10295
JournalScientific reports
Volume7
Issue number1
DOIs
StatePublished - Dec 1 2017

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Complementarity Determining Regions
Antibodies
Computer Simulation
Peptides
Antigens
Single-Chain Antibodies
Immunoglobulin Fab Fragments
Antibody Specificity
Amino Acid Substitution
Bacteriophages
Libraries
FLAG peptide
Epitopes
Immunization
Carrier Proteins
Clone Cells
Enzyme-Linked Immunosorbent Assay
Light
Sensitivity and Specificity

All Science Journal Classification (ASJC) codes

  • General

Cite this

Entzminger, K. C., Hyun, J. M., Pantazes, R. J., Patterson-Orazem, A. C., Qerqez, A. N., Frye, Z. P., ... Maynard, J. A. (2017). De novo design of antibody complementarity determining regions binding a FLAG tetra-peptide. Scientific reports, 7(1), [10295]. https://doi.org/10.1038/s41598-017-10737-9
Entzminger, Kevin C. ; Hyun, Jeong Min ; Pantazes, Robert J. ; Patterson-Orazem, Athena C. ; Qerqez, Ahlam N. ; Frye, Zach P. ; Hughes, Randall A. ; Ellington, Andrew D. ; Lieberman, Raquel L. ; Maranas, Costas D. ; Maynard, Jennifer A. / De novo design of antibody complementarity determining regions binding a FLAG tetra-peptide. In: Scientific reports. 2017 ; Vol. 7, No. 1.
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Entzminger, KC, Hyun, JM, Pantazes, RJ, Patterson-Orazem, AC, Qerqez, AN, Frye, ZP, Hughes, RA, Ellington, AD, Lieberman, RL, Maranas, CD & Maynard, JA 2017, 'De novo design of antibody complementarity determining regions binding a FLAG tetra-peptide', Scientific reports, vol. 7, no. 1, 10295. https://doi.org/10.1038/s41598-017-10737-9

De novo design of antibody complementarity determining regions binding a FLAG tetra-peptide. / Entzminger, Kevin C.; Hyun, Jeong Min; Pantazes, Robert J.; Patterson-Orazem, Athena C.; Qerqez, Ahlam N.; Frye, Zach P.; Hughes, Randall A.; Ellington, Andrew D.; Lieberman, Raquel L.; Maranas, Costas D.; Maynard, Jennifer A.

In: Scientific reports, Vol. 7, No. 1, 10295, 01.12.2017.

Research output: Contribution to journalArticle

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Entzminger KC, Hyun JM, Pantazes RJ, Patterson-Orazem AC, Qerqez AN, Frye ZP et al. De novo design of antibody complementarity determining regions binding a FLAG tetra-peptide. Scientific reports. 2017 Dec 1;7(1). 10295. https://doi.org/10.1038/s41598-017-10737-9