TY - JOUR
T1 - ITEM-THREE analysis of a monoclonal anti-malaria antibody reveals its assembled epitope on the pfMSP119 antigen
AU - Opuni, Kwabena F.M.
AU - Koy, Cornelia
AU - Russ, Manuela
AU - Reepmeyer, Maren
AU - Danquah, Bright D.
AU - Weresow, Moritz
AU - Alef, Astrid
AU - Lorenz, Peter
AU - Thiesen, Hans Juergen
AU - Glocker, Michael O.
N1 - Publisher Copyright:
© 2020 Opuni et al. Published under exclusive license by The American Society for Biochemistry and Molecular Biology, Inc.
PY - 2020/10/30
Y1 - 2020/10/30
N2 - 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 MBPpfMSP119 antigen surface that is recognized by the antipfMSP119 antibody G17.12. We identified three epitope-carrying peptides, 386GRNISQHQCVKKQCPQNSGCFRHLDE411386GRNISQHQCVKKQCPQNSGCFRHLDEREE414, and 415CKC-LLNYKQE424, 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.
AB - 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 MBPpfMSP119 antigen surface that is recognized by the antipfMSP119 antibody G17.12. We identified three epitope-carrying peptides, 386GRNISQHQCVKKQCPQNSGCFRHLDE411386GRNISQHQCVKKQCPQNSGCFRHLDEREE414, and 415CKC-LLNYKQE424, 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.
UR - http://www.scopus.com/inward/record.url?scp=85094983330&partnerID=8YFLogxK
U2 - 10.1074/jbc.RA120.014802
DO - 10.1074/jbc.RA120.014802
M3 - Article
C2 - 32848020
AN - SCOPUS:85094983330
SN - 0021-9258
VL - 295
SP - 14987
EP - 14997
JO - Journal of Biological Chemistry
JF - Journal of Biological Chemistry
IS - 44
ER -