ITEM-THREE analysis of a monoclonal anti-malaria antibody reveals its assembled epitope on the pfMSP119 antigen.
J Biol Chem
; 295(44): 14987-14997, 2020 10 30.
Article
em En
| MEDLINE
| ID: mdl-32848020
ABSTRACT
Rapid diagnostic tests are first-line assays for diagnosing infectious diseases, such as malaria. To minimize false positive and false negative test results in population-screening assays, high-quality reagents and well-characterized antigens and antibodies are needed. An important property of antigen-antibody binding is recognition specificity, which best can be estimated by mapping an antibody's epitope on the respective antigen. We have cloned a malarial antigen-containing fusion protein, MBP-pfMSP119, in Escherichia coli, which then was structurally and functionally characterized before and after high pressure-assisted enzymatic digestion. We then used our previously developed method, intact transition epitope mapping-targeted high-energy rupture of extracted epitopes (ITEM-THREE), to map the area on the MBP-pfMSP119 antigen surface that is recognized by the anti-pfMSP119 antibody G17.12. We identified three epitope-carrying peptides, 386GRNISQHQCVKKQCPQNSGCFRHLDE411, 386GRNISQHQCVKKQCPQNSGCFRHLDEREE414, and 415CKCLLNYKQE424, from the GluC-derived peptide mixture. These peptides belong to an assembled (conformational) epitope on the MBP-pfMSP119 antigen whose identification was substantiated by positive and negative control experiments. In conclusion, our data help to establish a workflow to obtain high-quality control data for diagnostic assays, including the use of ITEM-THREE as a powerful analytical tool. Data are available via ProteomeXchange PXD019717.
Palavras-chave
Texto completo:
1
Coleções:
01-internacional
Contexto em Saúde:
2_ODS3
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3_ND
Base de dados:
MEDLINE
Assunto principal:
Malária Falciparum
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Proteína 1 de Superfície de Merozoito
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Anticorpos Monoclonais
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Epitopos
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Antígenos de Protozoários
Limite:
Animals
Idioma:
En
Revista:
J Biol Chem
Ano de publicação:
2020
Tipo de documento:
Article