Encapsidated-CpG ODN enhances immunogenicity of porcine circovirus type 2 virus-like particles

Hansoongnern, Payuda; Phecharat, Nantawan; Wasanasuk, Ketkaew; Tommeurd, Wantanee; Chankeeree, Penpitcha; Lekcharoensuk, Chalermpol; Semkum, Ploypailin; Pinitkiatisakul, Sunan; Lekcharoensuk, Porntippa

doi:10.1016/j.vetmic.2022.109583

Cited by 2 publications

(3 citation statements)

References 46 publications

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“…Primary and secondary antibodies were ASFV convalescent swine serum at a dilution of 1:200 and anti-pig IgG HRP (Sigma) at a dilution of 1:1000, respectively. The bovine ephemeral fever virus G protein served as a negative control in this procedure [ 29 ].…”

Section: Methodsmentioning

confidence: 99%

Small ubiquitin-like modifier-tag and modified protein purification significantly increase the quality and quantity of recombinant African swine fever virus p30 protein

Chootip,

Hansoongnern,

Thangthamniyom

et al. 2024

Vet World

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Background and Aim: African swine fever (ASF) is a highly virulent and contagious viral disease caused by the ASF virus (ASFV). It has a significant impact on swine production throughout the world, while existing vaccines and specific treatments remain ineffective. ASFV p30 is a potent antigenic protein that induces protective antibodies immediately after infection; however, most recombinant p30 is insoluble. This study aimed to improve the solubility, yield, and purity of recombinant p30 by tagging it with a small ubiquitin-like modifier (SUMO) and modifying the protein purification process. Materials and Methods: SUMO fused with ASFV p30 (SUMO-p30) and p30 alone were cloned and expressed in Escherichia coli. SUMO-p30 and p30 solubility and expression levels were analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). Protein purification was modified by combining ammonium sulfate precipitation method with affinity chromatography. In addition, large-scale production of all versions of p30 were compared using SDS-PAGE and western blotting, and the purified p30 was used to develop the indirect enzyme-linked immunosorbent assay (ELISA). Results: The solubility and expression levels of SUMO-p30 were dramatically enhanced compared with that of p30. Modification of the purification process significantly increased purified and soluble SUMO-p30 and p30 yields by 6.59 and 1.02 μg/mL, respectively. Large-scale production confirmed that this procedure increased the quantity of recombinant p30 while maintaining protein purity and immunogenicity. The p30-based indirect ELISA was able to discriminate between positive and negative serum samples with statistically significant differences in mean optical density 450 values (p < 0.001). Conclusion: This study demonstrates the enhancement of solubility, purity, and yield of ASFV p30 expressed in E.coli by SUMO fusion tagging and combining ammonium sulfate precipitation with affinity chromatography for protein purification. These positive effects were sustained in large-scale production. Cleavage and removal of hexahistidine-SUMO tag from the fusion protein by protease may not be suitable when handling a large amount of the protein. However, the SUMO-fused p30 retained strong immunoreactivity to convalescent swine serum, indicating its application in immunization and diagnostic purposes. The expression and purification procedures in this study could be applied to increase solubility, quality, and quantity of other recombinant proteins as well. Keywords: African swine fever, diagnosis, indirect enzyme-linked immunosorbent assay, p30, protein expression, purification.

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Section: Methodsmentioning

confidence: 99%

Small ubiquitin-like modifier-tag and modified protein purification significantly increase the quality and quantity of recombinant African swine fever virus p30 protein

Chootip,

Hansoongnern,

Thangthamniyom

et al. 2024

Vet World

Self Cite

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“… 2007 ; Hansoongnern et al. 2022 ). The fact that incorporated tandem CpG-ODNs increase the PCV2 virus-like particle (VLP) vaccine efficacy underlines the importance of CpG motifs in modulation of cellular signals (Hansoongnern et al.…”

Section: Immunomodulatory Elements Of Circovirusesmentioning

confidence: 99%

“…The fact that incorporated tandem CpG-ODNs increase the PCV2 virus-like particle (VLP) vaccine efficacy underlines the importance of CpG motifs in modulation of cellular signals (Hansoongnern et al. 2022 ). In addition to the DNA sequence, the secondary and tertiary structure of the circular genomic DNA markedly influence the final action on the immune reaction (Wikstrom et al.…”

Section: Immunomodulatory Elements Of Circovirusesmentioning

confidence: 99%

Mechanisms of circovirus immunosuppression and pathogenesis with a focus on porcine circovirus 2: a review

Fehér

Jakab

Bányai

2023

Veterinary Quarterly

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Certain pathogens, due to their adverse effects on the immune reaction, aggravate the course of concomitant heterologous infections. Here we summarize mechanisms by which circoviruses, including the most studied porcine circovirus 2, and other mammalian and avian circoviruses, trigger their own replication and confound the hosts’ immune response. At different stages of infection, from latent state to disease induction, these viruses markedly influence the cellular signaling pathways. Circoviruses have been found to interfere with interferon and proinflammatory cytokine producing and responsive pathways. Apoptotic processes, altered cellular transport and constraint of the mitotic phase all support the viral replication. The cytokine imbalance and lymphocyte depletion, thus the impaired immunity, favors invasion of super- or co-infecting agents, which in concert with circoviruses induce illnesses with increased severity. The information summarized in this review point out the diversity of host and viral factors involved in the mechanisms of disease progression during circovirus infections.

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Encapsidated-CpG ODN enhances immunogenicity of porcine circovirus type 2 virus-like particles

Cited by 2 publications

References 46 publications

Small ubiquitin-like modifier-tag and modified protein purification significantly increase the quality and quantity of recombinant African swine fever virus p30 protein

Small ubiquitin-like modifier-tag and modified protein purification significantly increase the quality and quantity of recombinant African swine fever virus p30 protein

Mechanisms of circovirus immunosuppression and pathogenesis with a focus on porcine circovirus 2: a review

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