Expression and purification of recombinant nattokinase in Spodoptera frugiperda cells

@article{Li2007ExpressionAP,
  title={Expression and purification of recombinant nattokinase in Spodoptera frugiperda cells},
  author={Xiaoxiang Li and Xiaoli Wang and Shaoling Xiong and Jing Zhang and Litao Cai and Yanyan Yang},
  journal={Biotechnology Letters},
  year={2007},
  volume={29},
  pages={1459-1464}
}
A recombinant baculovirus, rv-egfp-NK, containing a reporter gene encoding the enhanced green fluorescent protein (EGFP), was used to express nattokinase (NK), a fibrinolytic enzyme, in Spodoptera frugiperda (SF-9) cells. The recombinant protein also included a histidine tag for purification using Ni2+ resins. The recombinant NK, ∼30 kDa, retained fibrinolytic activity (60 U/ml). The integration of the EGFP expression cassette in the Bac-to-Bac system is thus an effective method for the… 
View on Springer
ncbi.nlm.nih.gov
10 Citations
Background Citations
4
Methods Citations
4

10 Citations

Extrachromosomal expression of nat05 gene encoding an alkaline serine protease from Bacillus subtilis N05
TLDR
The R4 recombinant clone that was found to secrete nattokinase can be used as a source of recombinant protein that can be further purified for various therapeutic applications, or the strain could be use as a probiotic bacterium in functional foods.
Transient expression of optimized and synthesized nattokinase gene in melon (Cucumis melo L.) fruit by agroinfiltration
TLDR
Quantitative real-time reverse transcription PCR (qRT-PCR) analysis and fibrinolytic activity assays showed that the expression level of recombinant NK controlled by E8 promoter was higher than that controlled by 35S promoter.
Heterologous expression of nattokinase from B. subtilis natto using Pichia pastoris GS115 and assessment of its thrombolytic activity
TLDR
This study suggests that the P. pastoris expression system is an ideal platform for the large-scale, low-cost preparation of nattokinase.
Nattokinase: production and application
TLDR
Nattokinase (NK), also known as subtilisin NAT, is one of the most considerable extracellular enzymes produced by Bacillus subtilis natto and is regarded as a valuable dietary supplement or nutraceutical.
A simple and cost-saving approach to optimize the production of subtilisin NAT by submerged cultivation of Bacillus subtilis natto.
TLDR
The optimal conditions for subtilisin NAT production by submerged cultivation and three variables/three levels of response surface methodology (RSM) using various inoculum densities, glucose concentrations, and defatted soybean concentrations as the three variables were studied.
Microbial therapeutic enzymes: A promising area of biopharmaceuticals
Nattokinase: An Oral Antithrombotic Agent for the Prevention of Cardiovascular Disease
TLDR
Recombinant technology represents a promising approach for the production of NK with high purity for its use in antithrombotic applications and opportunities for utilizing plant systems for the large-scale production are discussed.
Recent Advances in Nattokinase-Enriched Fermented Soybean Foods: A Review
TLDR
Due to the continuous exploration and rapid progress of NK, NK is expected to be a natural future alternative to CVDs.
Fibrinolytic enzymes of Bacillus spp.: an overview.
TLDR
This review has made an attempt to present the importance of Bacillus spp.
Solubility, particle formation and immune display of trimers of major capsid protein 7 of African horsesickness virus fused with enhanced green fluorescent protein

References

SHOWING 1-10 OF 10 REFERENCES
Insect larval expression process is optimized by generating fusions with green fluorescent protein.
TLDR
The insect larvae/baculovirus protein production process was dramatically simplified by expressing fusion proteins containing green fluorescent protein (GFP) and the product-of-interest, and this technique obviates the need for in-process Western analyses.
Developing baculovirus-insect cell expression systems for humanized recombinant glycoprotein production.
Highly efficient gene expression of a fibrinolytic enzyme (subtilisin DFE) in Bacillus subtilis mediated by the promoter of α-amylase gene from Bacillus amyloliquefaciens
TLDR
Recombinant subtilisin DFE gene was successfully expressed in B. subtilis WB600 with a fibrinolytic activity of 200 urokinase units ml−1 and was inserted into the Escherichia coli/Bacillus subtilis shuttle plasmid vector, pSUGV4.
Efficient transduction of neural cells in vitro and in vivo by a baculovirus-derived vector.
TLDR
Baculovirus-derived vector, the Bac-CMV-GFP vector, containing a reporter gene encoding for the green fluorescent protein (GFP) under the control of the cytomegalovirus (CMV) promoter could be efficiently transduced and could transduce neural cells in vivo when directly injected into the brain of rodents.
Formation of Wild-Type and Chimeric Influenza Virus-Like Particles following Simultaneous Expression of Only Four Structural Proteins
TLDR
The described approach to assembly of wild-type and chimeric influenza VLPs may provide a valuable tool for further investigation of viral morphogenesis and genome packaging as well as for the development of novel vaccines.
Purification and characterization of a strong fibrinolytic enzyme (nattokinase) in the vegetable cheese natto, a popular soybean fermented food in Japan.
TLDR
The results indicate that nattokinase is a subtilisin-like serine protease, a strong fibrinolytic enzyme that digested not only fibrin but also several synthetic substrates.
Intracellular trafficking and interactions of the HIV-1 Tat protein.
Fusions of the human immunodeficiency virus type 1 (HIV-1) transactivator protein Tat to the green fluorescent protein (GFP) were used to study the intracellular localization, trafficking, and…
Partial purification of nattokinase from Bacillus subtilis by expanded bed adsorption
Nattokinase was purified from unclarified Bacillus subtilisculture filtrate using an expanded bed with a purification factor of 8.2 and at a yield of 95%. The optimal pHs for adsorption and elution…
A novel fibrinolytic enzyme (nattokinase) in the vegetable cheese Natto; a typical and popular soybean food in the Japanese diet
TLDR
A novel fibrinolytic enzyme, named nattokinase, was demonstrated in the vegetable cheese Natto, which is a typical soybean food eaten in Japan, and it digested fibrin but also the plasmin substrate H-D-Val-Leu-Lys-pNA, which was more sensitive to the enzyme than other substrates tried.
The fibrin plate method for estimating fibrinolytic activity.