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Heterologous Expression, Purification, and Immunomodulatory Effects of Recombinant Lipoprotein GUDIV-103 Isolated from Ureaplasma diversum

Ureaplasma diversum is a bacterial pathogen that infects cattle and can cause severe inflammation of the genital and reproductive systems. Lipid-associated membrane proteins (LAMPs), including GUDIV-103, are the main virulence factors in this bacterium. In this study, we heterologously expressed rec...

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Autores principales: Santos-Junior, Manoel Neres, Neves, Wanderson Souza, Santos, Ronaldo Silva, Almeida, Palloma Porto, Fernandes, Janaina Marinho, Guimarães, Bruna Carolina de Brito, Barbosa, Maysa Santos, da Silva, Lucas Santana Coelho, Gomes, Camila Pacheco, Sampaio, Beatriz Almeida, Rezende, Izadora de Souza, Correia, Thiago Macedo Lopes, Neres, Nayara Silva de Macedo, Campos, Guilherme Barreto, Bastos, Bruno Lopes, Timenetsky, Jorge, Marques, Lucas Miranda
Formato: Online Artículo Texto
Lenguaje:English
Publicado: MDPI 2022
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9147684/
https://www.ncbi.nlm.nih.gov/pubmed/35630474
http://dx.doi.org/10.3390/microorganisms10051032
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author Santos-Junior, Manoel Neres
Neves, Wanderson Souza
Santos, Ronaldo Silva
Almeida, Palloma Porto
Fernandes, Janaina Marinho
Guimarães, Bruna Carolina de Brito
Barbosa, Maysa Santos
da Silva, Lucas Santana Coelho
Gomes, Camila Pacheco
Sampaio, Beatriz Almeida
Rezende, Izadora de Souza
Correia, Thiago Macedo Lopes
Neres, Nayara Silva de Macedo
Campos, Guilherme Barreto
Bastos, Bruno Lopes
Timenetsky, Jorge
Marques, Lucas Miranda
author_facet Santos-Junior, Manoel Neres
Neves, Wanderson Souza
Santos, Ronaldo Silva
Almeida, Palloma Porto
Fernandes, Janaina Marinho
Guimarães, Bruna Carolina de Brito
Barbosa, Maysa Santos
da Silva, Lucas Santana Coelho
Gomes, Camila Pacheco
Sampaio, Beatriz Almeida
Rezende, Izadora de Souza
Correia, Thiago Macedo Lopes
Neres, Nayara Silva de Macedo
Campos, Guilherme Barreto
Bastos, Bruno Lopes
Timenetsky, Jorge
Marques, Lucas Miranda
author_sort Santos-Junior, Manoel Neres
collection PubMed
description Ureaplasma diversum is a bacterial pathogen that infects cattle and can cause severe inflammation of the genital and reproductive systems. Lipid-associated membrane proteins (LAMPs), including GUDIV-103, are the main virulence factors in this bacterium. In this study, we heterologously expressed recombinant GUDIV-103 (rGUDIV-103) in Escherichia coli, purified it, and evaluated its immunological reactivity and immunomodulatory effects in bovine peripheral blood mononuclear cells (PBMCs). Samples from rabbits inoculated with purified rGUDIV-103 were analysed using indirect enzyme-linked immunosorbent assay and dot blotting to confirm polyclonal antibody production and assess kinetics, respectively. The expression of this lipoprotein in field isolates was confirmed via Western blotting with anti-rGUDIV-103 serum and hydrophobic or hydrophilic proteins from 42 U. diversum strains. Moreover, the antibodies produced against the U. diversum ATCC 49783 strain recognised rGUDIV-103. The mitogenic potential of rGUDIV-103 was evaluated using a lymphoproliferation assay in 5(6)-carboxyfluorescein diacetate succinimidyl ester–labelled bovine PBMCs, where it induced lymphocyte proliferation. Quantitative polymerase chain reaction analysis revealed that the expression of interleukin-1β, toll-like receptor (TLR)-α, TLR2, TLR4, inducible nitric oxide synthase, and caspase-3–encoding genes increased more in rGUDIV-103–treated PBMCs than in untreated cells (p < 0.05). Treating PBMCs with rGUDIV-103 increased nitric oxide and hydrogen peroxide levels. The antigenic and immunogenic properties of rGUDIV-103 suggested its suitability for immunobiological application.
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spelling pubmed-91476842022-05-29 Heterologous Expression, Purification, and Immunomodulatory Effects of Recombinant Lipoprotein GUDIV-103 Isolated from Ureaplasma diversum Santos-Junior, Manoel Neres Neves, Wanderson Souza Santos, Ronaldo Silva Almeida, Palloma Porto Fernandes, Janaina Marinho Guimarães, Bruna Carolina de Brito Barbosa, Maysa Santos da Silva, Lucas Santana Coelho Gomes, Camila Pacheco Sampaio, Beatriz Almeida Rezende, Izadora de Souza Correia, Thiago Macedo Lopes Neres, Nayara Silva de Macedo Campos, Guilherme Barreto Bastos, Bruno Lopes Timenetsky, Jorge Marques, Lucas Miranda Microorganisms Article Ureaplasma diversum is a bacterial pathogen that infects cattle and can cause severe inflammation of the genital and reproductive systems. Lipid-associated membrane proteins (LAMPs), including GUDIV-103, are the main virulence factors in this bacterium. In this study, we heterologously expressed recombinant GUDIV-103 (rGUDIV-103) in Escherichia coli, purified it, and evaluated its immunological reactivity and immunomodulatory effects in bovine peripheral blood mononuclear cells (PBMCs). Samples from rabbits inoculated with purified rGUDIV-103 were analysed using indirect enzyme-linked immunosorbent assay and dot blotting to confirm polyclonal antibody production and assess kinetics, respectively. The expression of this lipoprotein in field isolates was confirmed via Western blotting with anti-rGUDIV-103 serum and hydrophobic or hydrophilic proteins from 42 U. diversum strains. Moreover, the antibodies produced against the U. diversum ATCC 49783 strain recognised rGUDIV-103. The mitogenic potential of rGUDIV-103 was evaluated using a lymphoproliferation assay in 5(6)-carboxyfluorescein diacetate succinimidyl ester–labelled bovine PBMCs, where it induced lymphocyte proliferation. Quantitative polymerase chain reaction analysis revealed that the expression of interleukin-1β, toll-like receptor (TLR)-α, TLR2, TLR4, inducible nitric oxide synthase, and caspase-3–encoding genes increased more in rGUDIV-103–treated PBMCs than in untreated cells (p < 0.05). Treating PBMCs with rGUDIV-103 increased nitric oxide and hydrogen peroxide levels. The antigenic and immunogenic properties of rGUDIV-103 suggested its suitability for immunobiological application. MDPI 2022-05-16 /pmc/articles/PMC9147684/ /pubmed/35630474 http://dx.doi.org/10.3390/microorganisms10051032 Text en © 2022 by the authors. https://creativecommons.org/licenses/by/4.0/Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https://creativecommons.org/licenses/by/4.0/).
spellingShingle Article
Santos-Junior, Manoel Neres
Neves, Wanderson Souza
Santos, Ronaldo Silva
Almeida, Palloma Porto
Fernandes, Janaina Marinho
Guimarães, Bruna Carolina de Brito
Barbosa, Maysa Santos
da Silva, Lucas Santana Coelho
Gomes, Camila Pacheco
Sampaio, Beatriz Almeida
Rezende, Izadora de Souza
Correia, Thiago Macedo Lopes
Neres, Nayara Silva de Macedo
Campos, Guilherme Barreto
Bastos, Bruno Lopes
Timenetsky, Jorge
Marques, Lucas Miranda
Heterologous Expression, Purification, and Immunomodulatory Effects of Recombinant Lipoprotein GUDIV-103 Isolated from Ureaplasma diversum
title Heterologous Expression, Purification, and Immunomodulatory Effects of Recombinant Lipoprotein GUDIV-103 Isolated from Ureaplasma diversum
title_full Heterologous Expression, Purification, and Immunomodulatory Effects of Recombinant Lipoprotein GUDIV-103 Isolated from Ureaplasma diversum
title_fullStr Heterologous Expression, Purification, and Immunomodulatory Effects of Recombinant Lipoprotein GUDIV-103 Isolated from Ureaplasma diversum
title_full_unstemmed Heterologous Expression, Purification, and Immunomodulatory Effects of Recombinant Lipoprotein GUDIV-103 Isolated from Ureaplasma diversum
title_short Heterologous Expression, Purification, and Immunomodulatory Effects of Recombinant Lipoprotein GUDIV-103 Isolated from Ureaplasma diversum
title_sort heterologous expression, purification, and immunomodulatory effects of recombinant lipoprotein gudiv-103 isolated from ureaplasma diversum
topic Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9147684/
https://www.ncbi.nlm.nih.gov/pubmed/35630474
http://dx.doi.org/10.3390/microorganisms10051032
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