Fructose 1,6-Bisphosphate Aldolase, a Novel Immunogenic Surface Protein on Listeria Species

Marcelo Mendonça; Gustavo Marçal Schmidt Garcia Moreira; Fabricio Rochedo Conceição; Michael Hust; Karla Sequeira Mendonça; Ângela Nunes Moreira; Rodrigo Correa França; Wladimir Padilha da Silva; Arun K Bhunia; José Antonio G Aleixo

doi:10.1371/journal.pone.0160544

Fructose 1,6-Bisphosphate Aldolase, a Novel Immunogenic Surface Protein on Listeria Species

PLoS One. 2016 Aug 4;11(8):e0160544. doi: 10.1371/journal.pone.0160544. eCollection 2016.

Affiliations

¹ Laboratório de Imunologia Aplicada, Núcleo de Biotecnologia, Centro de Desenvolvimento Tecnológico, Universidade Federal de Pelotas, Pelotas, RS, Brazil.
² Molecular Food Microbiology Laboratory, Department of Food Science, Purdue University, West Lafayette, Indiana, United Sates of America.
³ Technische Universität Braunschweig, Institut für Biochemie, Biotechnologie und Bioinformatik, Abteilung Biotechnologie, Spielmannstr, Braunschweig, Germany.
⁴ Laboratório de Microbiologia de Alimentos, Departamento de Ciência e Tecnologia Agroindustrial, Faculdade de Agronomia Eliseu Maciel, Universidade Federal de Pelotas, Pelotas, RS, Brazil.

Abstract

Listeria monocytogenes is a ubiquitous food-borne pathogen, and its presence in food or production facilities highlights the importance of surveillance. Increased understanding of the surface exposed antigens on Listeria would provide potential diagnostic and therapeutic targets. In the present work, using mass spectrometry and genetic cloning, we show that fructose-1,6-bisphosphate aldolase (FBA) class II in Listeria species is the antigen target of the previously described mAb-3F8. Western and dot blot assays confirmed that the mAb-3F8 could distinguish all tested Listeria species from close-related bacteria. Localization studies indicated that FBA is present in every fraction of Listeria cells, including supernatant and the cell wall, setting Listeria spp. as one of the few bacteria described to have this protein on their cell surface. Epitope mapping using ORFeome display and a peptide membrane revealed a 14-amino acid peptide as the potential mAb-3F8 epitope. The target epitope in FBA allowed distinguishing Listeria spp. from closely-related bacteria, and was identified as part of the active site in the dimeric enzyme. However, its function in cell surface seems not to be host cell adhesion-related. Western and dot blot assays further demonstrated that mAb-3F8 together with anti-InlA mAb-2D12 could differentiate pathogenic from non-pathogenic Listeria isolated from artificially contaminated cheese. In summary, we report FBA as a novel immunogenic surface target useful for the detection of Listeria genus.

MeSH terms

Amino Acid Sequence
Antibodies, Monoclonal / immunology
Antigens, Surface / immunology*
Antigens, Surface / metabolism
Bacterial Proteins / genetics
Bacterial Proteins / immunology*
Bacterial Proteins / metabolism
Blotting, Western
Catalytic Domain
Cheese / microbiology
Cloning, Molecular
Dimerization
Enzyme-Linked Immunosorbent Assay
Epitope Mapping
Epitopes / immunology
Food Microbiology
Fructose-Bisphosphate Aldolase / genetics
Fructose-Bisphosphate Aldolase / immunology*
Fructose-Bisphosphate Aldolase / metabolism
Listeria / enzymology*
Listeria / immunology*
Listeria / isolation & purification
Mass Spectrometry
Peptides / analysis
Protein Structure, Tertiary
Recombinant Proteins / biosynthesis
Recombinant Proteins / immunology
Recombinant Proteins / isolation & purification

Substances

Antibodies, Monoclonal
Antigens, Surface
Bacterial Proteins
Epitopes
Peptides
Recombinant Proteins
Fructose-Bisphosphate Aldolase

Grants and funding

The authors thank Coordenadoria de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) and Fundação de Amparo à Pesquisa do Estado do Rio Grande do Sul (FAPERGS) for the scholarship of Marcelo Mendonça and the Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) from Brazil for scholarships to Gustavo Moriera and financial support (Project numbers 483673/2013-7, and 204693/2014-4); the agricultural Research Service of the U.S. Department of Agriculture (Project number 1935-42000-072-02G); and the Center for Food Safety and Engineering at Purdue University for hosting Marcelo Mendonça as a visiting scientist for sandwich PhD program and for the financial support to the work. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.