Recombinant expression and functional analysis of antimicrobial Siganus oramin l-amino acid oxidase using the Bac-to-Bac baculovirus expression system

Han, Xiao; Huang, Yuxi; Hou, Yulin; Dang, Huifeng; Li, Ruijun

doi:10.1016/j.fsi.2019.11.062

Cited by 9 publications

(4 citation statements)

References 19 publications

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“…The bac-to-bac baculovirus expression system is one of them, which has advantages of post-translation modification, high biological activity, and efficient expression of foreign genes [ 19 , 20 ]. Furthermore, the baculovirus cannot infect mammals; therefore, it is not a safety concern to humans [ 21 ]. In light of these advantages, a bac-to-bac expression system was employed to generate GCRV genotype II based VLPs.…”

Section: Introductionmentioning

confidence: 99%

Recombinant Baculovirus-Produced Grass Carp Reovirus Virus-Like Particles as Vaccine Candidate That Provides Protective Immunity against GCRV Genotype II Infection in Grass Carp

Gao

et al. 2021

Vaccines

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Grass carp reovirus (GCRV) leads to severe hemorrhagic disease in grass carp (Ctenopharyngodon idella) and causes economic losses in grass carp aquaculture. Recent epidemiological investigations showed that GCRV genotype II is the dominant subtype in China. Therefore, it is very important to develop a novel vaccine for preventing diseases caused by GCRV genotype II. In this study, we employed a bac-to-bac expression system to generate GCRV-II-based virus-like particles (VLPs). Previous studies have shown that the structural proteins VP3, VP4, and VP38 encoded by the segments S3, S6, and S10 of type II GCRV are immunogenic. Hence, the GCRV-VLPs were produced by co-infection of sf9 cells with recombinant baculoviruses PFBH-VP3, PFBH-VP4, and PFBH-VP38. The expressions of VP3, VP4, and VP38 proteins in GCRV-VLPs were tested by IFA and Western blot analysis. By electron microscopic observations of ultrathin sections, purified VLPs showed that the expressed proteins are similar in shape to GCRV genotype II with a size range from 40 nm to 60 nm. The immunogenicity of GCRV-VLPs was evaluated by the injection immunization of grass carp. The analysis of serum-specific IgM antibody showed that grass carp immunized with GCRV-VLPs produced GCRV-specific antibodies. Furthermore, injection with GCRV-VLPs increased the expressions of immune-related genes (IgM, IFN, TLR3, TLR7) in the spleen and kidney. In addition, grass carp immunized with a GCRV-VLPs-based vaccine showed a relative percent survival rate (RPS) of 83.33% after challenge. The data in this study showed that GCRV-VLPs demonstrated an excellent immunogenicity and represent a promising approach for vaccine development against GCRV genotype II infection.

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Section: Introductionmentioning

confidence: 99%

Recombinant Baculovirus-Produced Grass Carp Reovirus Virus-Like Particles as Vaccine Candidate That Provides Protective Immunity against GCRV Genotype II Infection in Grass Carp

Gao

et al. 2021

Vaccines

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“…The expression vectors of TcFOXO and TcCncC were constructed by cloning the PCR products into the linearized pFastBacHT B expression vector with the restriction enzymes KpnI and BamHI, and the fusions were performed with an In‐Fusion HD cloning kit. The pFastBac–TcFOXO and pFastBac–TcCncC plasmids were subsequently transformed into the DH10Bac competent cells, and the recombinant bacmids of Bacmid–TcFOXO and Bacmid–TcCncC were extracted by using a plasmid extraction kit (TIANGEN) according to the manufacturer’s instructions 47 . The transfections of bacmids into Sf9 cells were performed as described above.…”

Section: Methodsmentioning

confidence: 99%

“…The pFastBac-TcFOXO and pFastBac-TcCncC plasmids were subsequently transformed into the DH10Bac competent cells, and the recombinant bacmids of Bacmid-TcFOXO and Bacmid-TcCncC were extracted by using a plasmid extraction kit (TIANGEN) according to the manufacturer's instructions. 47 The transfections of bacmids into Sf9 cells were performed as described above. The supernatant was collected after 48 h of transfection, and 200 μl of the supernatant was added into 2 ml of Sf-900 II SFM in six-well plates.…”

Section: Electrophoretic Mobility Shift Assaysmentioning

confidence: 99%

FOXO acts as a positive regulator of CncC and deltamethrin tolerance in the red flour beetle, Tribolium castaneum

Zhang

Meng

Jiang

et al. 2022

Pest Management Science

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BACKGROUND Both forkhead box O (FOXO) and nuclear factor erythroid‐derived 2‐like‐2 (Nrf2) are key transcription factors related to stress responses. Whereas limited studies in mammals and Caenorhabditis elegans have revealed the interaction between FoxO/DAF‐16 and Nrf2/SKN‐1, the role of FOXO in metabolic detoxification and regulation of the Nrf2–Keap1 signaling pathway are poorly understood in insects. RESULTS Using Tribolium castaneum as a model organism, we found that RNA interference‐mediated knockdown of FOXO enhanced deltamethrin‐induced lethality by affecting the messenger RNA (mRNA) levels of CYP6BQ cluster genes. We further demonstrated that injection of dsFOXO into the beetle larvae decreased expression of CncC and KEAP1 at both the mRNA and protein level. Notably, dual‐luciferase and electrophoretic mobility shift assays both confirmed direct regulation of CncC by FOXO, whereas Keap1 was directly regulated by CncC. CONCLUSION FOXO can directly regulate the expression of CncC and indirectly regulate the expression of Keap1 through CncC. The data provide insights into the regulatory mechanisms of the Nrf2–Keap1 signaling pathway in insects.

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“…Further, hcLAAO4, one of the fungal LAAOs from Hebeloma cylindrosporum , was expressed in E. coli or yeast ( Pichia pastors ) systems with His-Tag (Bloess et al 2019 ; Heß et al 2020 ). Moreover, L-Lys-specific LAAO isolated from the Siganus pramin (serum) or Platichthys stellatus (gill), which is an innate immune protein with antibacterial activity, was codon optimized for expression in insect cells (Sf9) or P. pastoris expression systems (Han et al 2020 , Kasai et al 2020 (Table 1 V1)). These recombinant LAAOs can be easily high mass-produced by scale-up and display strong bioactivity via bactericidal, parasiticidal, and apoptotic effects, and bind to the surface of bacteria, similar to native LAAOs, suggesting potential use of LAAOs against clinical pathogens.…”

Section: Perspectives For Possible Biomedical Applicationsmentioning

confidence: 99%

Antimicrobial properties of L-amino acid oxidase: biochemical features and biomedical applications

Kasai

Nakano

Ohishi

et al. 2021

Appl Microbiol Biotechnol

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Mucus layer that covers the body surface of various animal functions as a defense barrier against microbes, environmental xenobiotics, and predators. Previous studies have reported that L-amino acid oxidase (LAAO), present in several animal fluids, has potent properties against pathogenic bacteria, viruses, and parasites. LAAO catalyzes the oxidative deamination of specific L-amino acids with the generation of hydrogen peroxide and L-amino acid metabolites. Further, the generated hydrogen peroxide is involved in oxidation (direct effect) while the metabolites activate immune responses (indirect effect). Therefore, LAAO exhibits two different mechanisms of bioactivation. Previously, we described the selective, specific, and local oxidative and potent antibacterial actions of various LAAOs as potential therapeutic strategies. In this review, we focus on their biochemical features, enzymatic regulations, and biomedical applications with a view of describing their probable role as biochemical agents and biomarkers for microbial infections, cancer, and autoimmune-mediated diseases. We consider that LAAOs hold implications in biomedicine owing to their antimicrobial activity wherein they can be used in treatment of infectious diseases and as diagnostic biomarkers in the above-mentioned diseased conditions. Key points •Focus on biochemical features, enzymatic regulation, and biomedical applications of LAAOs. •Mechanisms of antimicrobial activity, inflammatory regulation, and immune responses of LAAOs. •Potential biomedical application as an antimicrobial and anti-infection agent, and disease biomarker.

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Recombinant expression and functional analysis of antimicrobial Siganus oramin l-amino acid oxidase using the Bac-to-Bac baculovirus expression system

Cited by 9 publications

References 19 publications

Recombinant Baculovirus-Produced Grass Carp Reovirus Virus-Like Particles as Vaccine Candidate That Provides Protective Immunity against GCRV Genotype II Infection in Grass Carp

Recombinant Baculovirus-Produced Grass Carp Reovirus Virus-Like Particles as Vaccine Candidate That Provides Protective Immunity against GCRV Genotype II Infection in Grass Carp

FOXO acts as a positive regulator of CncC and deltamethrin tolerance in the red flour beetle, Tribolium castaneum

Antimicrobial properties of L-amino acid oxidase: biochemical features and biomedical applications

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