Construction and expression of a heterologous protein in Lactococcus lactis by using the nisin-controlled gene expression system: the case of the PRRSV ORF6 gene

Wang, Z.H.; Wang, Y.L.; Zeng, Xianyin

doi:10.4238/2014.february.20.10

Cited by 8 publications

(7 citation statements)

References 27 publications

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“…Moreover, in the mechanism of action known as detergent, peptides can cause rupture of the layers and consequent loss of ions ( Wimley, 2010 ). Some studies consider the action of AMPs on enzymes, gene transcription and DNA synthesis ( Wang et al, 2014 ). To date, there have been over 5000 AMPs reported ( Bahar and Ren, 2013 ), with different ways of acting.…”

Section: Introduction To Antimicrobial Peptides and Their Importancementioning

confidence: 99%

“…Peptide design allows amino acid residue modifications, increasing their antimicrobial and immunomodulatory capacity, or even improving their resistance against degradation ( Riedl et al, 2011 ). Synthetic peptides can also be expressed in other organisms through heterologous expression techniques, allowing a reduction in price when produced on a large scale ( Wang et al, 2014 ).…”

Section: Introduction To Antimicrobial Peptides and Their Importancementioning

confidence: 99%

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Antimicrobial Peptides and Nanotechnology, Recent Advances and Challenges

et al. 2018

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Antimicrobial peptides are sequences of amino acids, which present activity against microorganisms. These peptides were discovered over 70 years ago, and are abundant in nature from soil bacteria, insects, amphibians to mammals and plants. They vary in amino acids number, the distance between amino acids within individual peptide structure, net charge, solubility and other physical chemical properties as well as differ in mechanism of action. These peptides may provide an alternative treatment to conventional antibiotics, which encounter resistance such as the peptide nisin applied in treating methicillin resistant Staphylococcus aureus (MRSA) or may behave synergistically with known antibiotics against parasites for instance, nisin Z when used in synergy with ampicillin reported better activity against Pseudomonas fluorescens than when the antibiotic was alone. AMPs are known to be active against viruses, bacteria, fungi and protozoans. Nanotechnology is an arena which explores the synthesis, characterization and application of an array of delivery systems at a one billionth of meter scale. Such systems are implemented to deliver drugs, proteins, vaccines, and peptides. The role of nanotechnology in delivering AMPs is still at its early development stage. There are challenges of incorporating AMPs into drug delivery system. This review intends to explore in depth, the role of nanotechnology in delivering AMPs as well as presenting the current advances and accompanying challenges of the technology.

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Section: Introduction To Antimicrobial Peptides and Their Importancementioning

confidence: 99%

Section: Introduction To Antimicrobial Peptides and Their Importancementioning

confidence: 99%

Antimicrobial Peptides and Nanotechnology, Recent Advances and Challenges

et al. 2018

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“…This host strain is a food grade and a progeny of NZ3000, which was developed for food grade application. This strain had been reported to be used for clinical test approach, secreting therapeutic protein, such as PRRSV [39]. Since this strain has a deletion of lacF, which is operon lactose integrated into the chromosome, for the expression level to depend on the inducer concentration.…”

Section: Discussionmentioning

confidence: 99%

Heterologous Expression of Interferon α-2b in Lactococcus lactis and its Biological Activity against Colorectal Cancer Cells

Meilina¹,

Budiarti²,

Mustopa³

et al. 2021

Kor. J. Microbiol. Biotechnol

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Type I Interferons (IFNα) are known for their role as biological anticancer agents owing to their cell-apoptosis inducing properties. Development of an appropriate, cost-effective host expression system is crucial for meeting the increasing demand for proteins. Therefore, this study aims to develop codon-optimized IFNα-2b in L. lactis NZ3900. These cells express extracellular protein using the NICE system and Usp 45 signal peptide. To validate the mature form of the expressed protein, the recombinant IFNα-2b was screened in a human colorectal cancer cell line using the cytotoxicity assay. The IFNα-2b was successfully cloned into the pNZ8148 vector, thereby generating recombinant L. lactis pNZ8148-SP Usp45 -IFNα-2b. The computational analysis of codon-optimized IFNα-2b revealed no mutation and amino acid changes; additionally, the codon-optimized IFNα-2b showed 100% similarity with native human IFNα-2b, in the BLAST analysis. The partial size exclusion chromatography (SEC) of extracellular protein yielded a 19 kDa protein, which was further confirmed by its positive binding to anti-IFNα-2b in the western blot analysis. The crude protein and SEC-purified partial fraction showed IC 50 values of 33.22 μg/ml and 127.2 μg/ml, respectively, which indicated better activity than the metabolites of L. lactis NZ3900 (231.8 μg/ml). These values were also comparable with those of the regular anticancer drug tamoxifen (105.5 μg/ml). These results demonstrated L. lactis as a promising host system that functions by utilizing the pNZ8148 NICE system. Meanwhile, codonoptimized usage of the inserted gene increased the optimal protein expression levels, which could be beneficial for its large-scale production. Taken together, the recombinant L. lactis IFNα-2b is a potential alternative treatment for colorectal cancer. Furthermore, its activity was analyzed in the WiDr cell line, to assess its colorectal anticancer activities in vivo.

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“…The results of this response will form a transcription factor that activates the promoter nisA, so that the transcription process by RNA polymerase can be functioning. 22 The presence of target protein bands in noninduced samples is expected due to basal activity causing the usp45-MSP-1 19 gene to be expressed even in the absence of nisin induction. This has been demonstrated in the Mohseni et al 21 study that NICE expression systems often exhibit basal expression activity (in the absence of induction) when expressed in Lactobacillus.…”

Section: Discussionmentioning

confidence: 99%

The production of SPusp45-MSP-1<sub>19</sub> gene construct and its recombinant protein in <em>Lactococcus lactis</em> to be used as a malaria vaccine

et al. 2018

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Background: Merozoite surface protein 1 (MSP-1) is a major protein used by the Plasmodium during red blood cells invasion in malaria. MSP-119, one of MSP-1 is highly conserved, and it is a potential malaria vaccine candidate because the monoclonal antibodies are capable blocking erythrocyte invasion in vitro. The aim of this study was to produce MSP-119 gene construct and the recombinant protein in Lactococcus lactis.Methods: Usp45-MSP-119, derived from codon optimization and the synthetic gene, was inserted into the pMAT cloning vector. A vector expressing MSP-119 included usp45 has been constructed by the manipulation of recombinant DNA using restriction enzymes. The MSP-119 protein was expressed to 45% ammonium sulfate precipitation and purified using Sephadex-G50 gel filtration chromatography. The expressed protein was characterized by SDS-PAGE and dot blot.Results: usp45-MSP-119 gene was amplified using specific primers and inserted into the multiple cloning sites in the expression vector pNZ8148 with size 3,538 bp as a recombinant vector. The protein of MSP-119 was successfully expressed in L. lactis with molecular weight of 10.45 kDa. The dot blot was tested in 3 different comparisons between the host cells, non-induced cells, and induced cells with 10 ng/ml nisin. The results showed that 10 ng/ml nisin gave a positive reaction as detected by dot blot assay.Conclusion: This study confirmed that the usp45-MSP-119 gene was successfully inserted into the multiple cloning sites of the pNZ8148 expression vector and the MSP-119 protein expressed in the NICE system of the L. lactis host cell.

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Construction and expression of a heterologous protein in Lactococcus lactis by using the nisin-controlled gene expression system: the case of the PRRSV ORF6 gene

Cited by 8 publications

References 27 publications

Antimicrobial Peptides and Nanotechnology, Recent Advances and Challenges

Antimicrobial Peptides and Nanotechnology, Recent Advances and Challenges

Heterologous Expression of Interferon α-2b in Lactococcus lactis and its Biological Activity against Colorectal Cancer Cells

The production of SPusp45-MSP-1<sub>19</sub> gene construct and its recombinant protein in <em>Lactococcus lactis</em> to be used as a malaria vaccine

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