Rapid fine conformational epitope mapping using comprehensive mutagenesis and deep sequencing

Caitlin A. Kowalsky, Matthew S. Faber, Aritro Nath, Hailey E. Dann, Vince W. Kelly, Li Liu, Purva Shanker, Ellen K. Wagner, Jennifer A. Maynard, Christina Chan, Timothy A. Whitehead

    Research output: Contribution to journalArticle

    • 11 Citations

    Abstract

    Knowledge of the fine location of neutralizing and non-neutralizing epitopes on human pathogens affords a better understanding of the structural basis of antibody efficacy, which will expedite rational design of vaccines, prophylactics, and therapeutics. However, full utilization of the wealth of information from single cell techniques and antibody repertoire sequencing awaits the development of a high throughput, inexpensive method to map the conformational epitopes for antibody-antigen interactions. Herewe show such an approach that combines comprehensive mutagenesis, cell surface display, and DNA deep sequencing. We develop analytical equations to identify epitope positions and show the method effectiveness by mapping the fine epitope for different antibodies targeting TNF, pertussis toxin, and the cancer target TROP2. Inall threecases, the experimentally determined conformational epitope was consistent with previous experimental datasets, confirming the reliability of the experimental pipeline. Once the comprehensive library is generated, fine conformational epitope maps canbe prepared at a rate of four per day.

    Original languageEnglish (US)
    Pages (from-to)26457-26470
    Number of pages14
    JournalJournal of Biological Chemistry
    Volume290
    Issue number44
    DOIs
    StatePublished - Oct 30 2015

    Profile

    Epitope Mapping
    High-Throughput Nucleotide Sequencing
    Mutagenesis
    Epitopes
    Antibodies
    Midodrine
    Hereditary Corneal Dystrophies
    Insulin Infusion Systems
    Pertussis Toxin
    DNA Sequence Analysis
    Vaccines
    Antigens
    Neoplasms
    Datasets
    Edema Disease of Swine
    Argininosuccinic Acid
    Breast Cyst
    Gossypium
    Aminopyrine
    Pathogens

    ASJC Scopus subject areas

    • Biochemistry
    • Cell Biology
    • Molecular Biology

    Cite this

    Kowalsky, C. A., Faber, M. S., Nath, A., Dann, H. E., Kelly, V. W., Liu, L., ... Whitehead, T. A. (2015). Rapid fine conformational epitope mapping using comprehensive mutagenesis and deep sequencing. Journal of Biological Chemistry, 290(44), 26457-26470. DOI: 10.1074/jbc.M115.676635

    Rapid fine conformational epitope mapping using comprehensive mutagenesis and deep sequencing. / Kowalsky, Caitlin A.; Faber, Matthew S.; Nath, Aritro; Dann, Hailey E.; Kelly, Vince W.; Liu, Li; Shanker, Purva; Wagner, Ellen K.; Maynard, Jennifer A.; Chan, Christina; Whitehead, Timothy A.

    In: Journal of Biological Chemistry, Vol. 290, No. 44, 30.10.2015, p. 26457-26470.

    Research output: Contribution to journalArticle

    Kowalsky, CA, Faber, MS, Nath, A, Dann, HE, Kelly, VW, Liu, L, Shanker, P, Wagner, EK, Maynard, JA, Chan, C & Whitehead, TA 2015, 'Rapid fine conformational epitope mapping using comprehensive mutagenesis and deep sequencing' Journal of Biological Chemistry, vol 290, no. 44, pp. 26457-26470. DOI: 10.1074/jbc.M115.676635
    Kowalsky CA, Faber MS, Nath A, Dann HE, Kelly VW, Liu L et al. Rapid fine conformational epitope mapping using comprehensive mutagenesis and deep sequencing. Journal of Biological Chemistry. 2015 Oct 30;290(44):26457-26470. Available from, DOI: 10.1074/jbc.M115.676635

    Kowalsky, Caitlin A.; Faber, Matthew S.; Nath, Aritro; Dann, Hailey E.; Kelly, Vince W.; Liu, Li; Shanker, Purva; Wagner, Ellen K.; Maynard, Jennifer A.; Chan, Christina; Whitehead, Timothy A. / Rapid fine conformational epitope mapping using comprehensive mutagenesis and deep sequencing.

    In: Journal of Biological Chemistry, Vol. 290, No. 44, 30.10.2015, p. 26457-26470.

    Research output: Contribution to journalArticle

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