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Development of Phospholipase A 2 Receptor Antigen Binding Monobodies for the Inhibition of the Idiopathic Membranous Glomerulonephritis Auto‐antibody Binding
Author(s) -
Stoddard Shana V,
Zaravar AliReza,
May Xavier,
Goldman Liam
Publication year - 2017
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.31.1_supplement.922.4
Subject(s) - epitope , antigen , in silico , docking (animal) , mutant , mutagenesis , binding selectivity , monoclonal antibody , chemistry , antibody , epitope mapping , biology , immunology , medicine , biochemistry , gene , nursing
The use of non‐specific treatment options in autoimmune therapy today unfortunately causes patients to be prone to other infections. This non‐specific therapy suppresses the entire patient's immunes system also making it harder to fight of basic infections. Thus therapies need to be developed which can accurately target only the aberrant portion of the immune system that contributes to the autoimmune disorder (AD) specific to the individual. The work here focuses on design of antigen specific binding monobodies as a targeted means to treat AD, and specifically idiopathic membranous glomerulonephritis. Sixty monobodies were designed and optimized using in silico mutagenesis. Protein‐protein docking simulations were performed using Rosetta to evaluate for maximum binding efficiency against the ricin and the CTLD2 epitopes present on the PLA 2 R antigen. Our results show improved binding scores and selectivity of monobodies against the phospholipase A 2 receptor (PLA 2 R) antigens. Initial binding energy, measured in rosetta energy units (REU), were −2.863 and −2.633 for the SH13 template and ySMB‐1 templates respectively. The M005 mutation designed in the SH13 improved binding scores to −5.32 REU. The M006 ySMB‐1 mutant showed the most in silico selectivity between the two epitope regions on the PLA 2 R antigen having a score of −2.966 REU against the CTLD2 epitope and a better score of −3.736 against the ricin epitope. The SH13 M005 mutant showed the greatest binding efficiency having a score of −5.319 REU. However, this mutant also binds the CTLD2 epitope (binding energy score produced was −5.32 REU), suggesting that it will not have selectivity. Several molecular interactions were identified that facilitated increased binding of the monobodies to the epitope regions compared to the original template monobodies. Molecular interactions leading to selectivity are also being evaluated. Seven monobody candidates were selected for expression which was performed in E. coli followed by subsequent purification using affinity chromatography for experimental verification of computational results is using ELISA. This work has implications for new antigen specific approach to AD therapy. Support or Funding Information William Randolph Hearst Foundation and Rhodes College Department of Chemistry