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Pyruvate dehydrogenase complex—enzyme 2, a new target for Listeria spp. detection identified using combined phage display technologies

The genus Listeria comprises ubiquitous bacteria, commonly present in foods and food production facilities. In this study, three different phage display technologies were employed to discover targets, and to generate and characterize novel antibodies against Listeria: antibody display for biomarker...

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Autores principales: Moreira, Gustavo Marçal Schmidt Garcia, Köllner, Sarah Mara Stella, Helmsing, Saskia, Jänsch, Lothar, Meier, Anja, Gronow, Sabine, Boedeker, Christian, Dübel, Stefan, Mendonça, Marcelo, Moreira, Ângela Nunes, Conceição, Fabricio Rochedo, Hust, Michael
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Nature Publishing Group UK 2020
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7498459/
https://www.ncbi.nlm.nih.gov/pubmed/32943681
http://dx.doi.org/10.1038/s41598-020-72159-4
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author Moreira, Gustavo Marçal Schmidt Garcia
Köllner, Sarah Mara Stella
Helmsing, Saskia
Jänsch, Lothar
Meier, Anja
Gronow, Sabine
Boedeker, Christian
Dübel, Stefan
Mendonça, Marcelo
Moreira, Ângela Nunes
Conceição, Fabricio Rochedo
Hust, Michael
author_facet Moreira, Gustavo Marçal Schmidt Garcia
Köllner, Sarah Mara Stella
Helmsing, Saskia
Jänsch, Lothar
Meier, Anja
Gronow, Sabine
Boedeker, Christian
Dübel, Stefan
Mendonça, Marcelo
Moreira, Ângela Nunes
Conceição, Fabricio Rochedo
Hust, Michael
author_sort Moreira, Gustavo Marçal Schmidt Garcia
collection PubMed
description The genus Listeria comprises ubiquitous bacteria, commonly present in foods and food production facilities. In this study, three different phage display technologies were employed to discover targets, and to generate and characterize novel antibodies against Listeria: antibody display for biomarker discovery and antibody generation; ORFeome display for target identification; and single-gene display for epitope characterization. With this approach, pyruvate dehydrogenase complex—enzyme 2 (PDC-E2) was defined as a new detection target for Listeria, as confirmed by immunomagnetic separation-mass spectrometry (IMS-MS). Immunoblot and fluorescence microscopy showed that this protein is accessible on the bacterial cell surface of living cells. Recombinant PDC-E2 was produced in E. coli and used to generate 16 additional antibodies. The resulting set of 20 monoclonal scFv-Fc was tested in indirect ELISA against 17 Listeria and 16 non-Listeria species. Two of them provided 100% sensitivity (CI 82.35–100.0%) and specificity (CI 78.20–100.0%), confirming PDC-E2 as a suitable target for the detection of Listeria. The binding region of 18 of these antibodies was analyzed, revealing that ≈ 90% (16/18) bind to the lipoyl domains (LD) of the target. The novel target PDC-E2 and highly specific antibodies against it offer new opportunities to improve the detection of Listeria.
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spelling pubmed-74984592020-09-18 Pyruvate dehydrogenase complex—enzyme 2, a new target for Listeria spp. detection identified using combined phage display technologies Moreira, Gustavo Marçal Schmidt Garcia Köllner, Sarah Mara Stella Helmsing, Saskia Jänsch, Lothar Meier, Anja Gronow, Sabine Boedeker, Christian Dübel, Stefan Mendonça, Marcelo Moreira, Ângela Nunes Conceição, Fabricio Rochedo Hust, Michael Sci Rep Article The genus Listeria comprises ubiquitous bacteria, commonly present in foods and food production facilities. In this study, three different phage display technologies were employed to discover targets, and to generate and characterize novel antibodies against Listeria: antibody display for biomarker discovery and antibody generation; ORFeome display for target identification; and single-gene display for epitope characterization. With this approach, pyruvate dehydrogenase complex—enzyme 2 (PDC-E2) was defined as a new detection target for Listeria, as confirmed by immunomagnetic separation-mass spectrometry (IMS-MS). Immunoblot and fluorescence microscopy showed that this protein is accessible on the bacterial cell surface of living cells. Recombinant PDC-E2 was produced in E. coli and used to generate 16 additional antibodies. The resulting set of 20 monoclonal scFv-Fc was tested in indirect ELISA against 17 Listeria and 16 non-Listeria species. Two of them provided 100% sensitivity (CI 82.35–100.0%) and specificity (CI 78.20–100.0%), confirming PDC-E2 as a suitable target for the detection of Listeria. The binding region of 18 of these antibodies was analyzed, revealing that ≈ 90% (16/18) bind to the lipoyl domains (LD) of the target. The novel target PDC-E2 and highly specific antibodies against it offer new opportunities to improve the detection of Listeria. Nature Publishing Group UK 2020-09-17 /pmc/articles/PMC7498459/ /pubmed/32943681 http://dx.doi.org/10.1038/s41598-020-72159-4 Text en © The Author(s) 2020 Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article's Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/.
spellingShingle Article
Moreira, Gustavo Marçal Schmidt Garcia
Köllner, Sarah Mara Stella
Helmsing, Saskia
Jänsch, Lothar
Meier, Anja
Gronow, Sabine
Boedeker, Christian
Dübel, Stefan
Mendonça, Marcelo
Moreira, Ângela Nunes
Conceição, Fabricio Rochedo
Hust, Michael
Pyruvate dehydrogenase complex—enzyme 2, a new target for Listeria spp. detection identified using combined phage display technologies
title Pyruvate dehydrogenase complex—enzyme 2, a new target for Listeria spp. detection identified using combined phage display technologies
title_full Pyruvate dehydrogenase complex—enzyme 2, a new target for Listeria spp. detection identified using combined phage display technologies
title_fullStr Pyruvate dehydrogenase complex—enzyme 2, a new target for Listeria spp. detection identified using combined phage display technologies
title_full_unstemmed Pyruvate dehydrogenase complex—enzyme 2, a new target for Listeria spp. detection identified using combined phage display technologies
title_short Pyruvate dehydrogenase complex—enzyme 2, a new target for Listeria spp. detection identified using combined phage display technologies
title_sort pyruvate dehydrogenase complex—enzyme 2, a new target for listeria spp. detection identified using combined phage display technologies
topic Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7498459/
https://www.ncbi.nlm.nih.gov/pubmed/32943681
http://dx.doi.org/10.1038/s41598-020-72159-4
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