BioMed Research International

BioMed Research International / 2014 / Article
Special Issue

Plant-Made Biologics

View this Special Issue

Review Article | Open Access

Volume 2014 |Article ID 249519 |

Hyun-Soon Kim, Jae-Heung Jeon, Kyung Jin Lee, Kisung Ko, "N-Glycosylation Modification of Plant-Derived Virus-Like Particles: An Application in Vaccines", BioMed Research International, vol. 2014, Article ID 249519, 8 pages, 2014.

N-Glycosylation Modification of Plant-Derived Virus-Like Particles: An Application in Vaccines

Academic Editor: Luca Santi
Received05 Feb 2014
Revised19 Apr 2014
Accepted12 May 2014
Published25 May 2014


Plants have been developed as an alternative system to mammalian cells for production of recombinant prophylactic or therapeutic proteins for human and animal use. Effective plant expression systems for recombinant proteins have been established with the optimal combination of gene expression regulatory elements and control of posttranslational processing of recombinant glycoproteins. In plant, virus-like particles (VLPs), viral “empty shells” which maintain the same structural characteristics of virions but are genome-free, are considered extremely promising as vaccine platforms and therapeutic delivery systems. Unlike microbial fermentation, plants are capable of carrying out N-glycosylation as a posttranslational modification of glycoproteins. Recent advances in the glycoengineering in plant allow human-like glycomodification and optimization of desired glycan structures for enhancing safety and functionality of recombinant pharmaceutical glycoproteins. In this review, the current plant-derived VLP approaches are focused, and N-glycosylation and its in planta modifications are discussed.

1. Plant-Derived Virus-Like Particle (VLP)

Viruses are able to form the quaternary structure of viral capsids through molecular self-assembly of repetitive building blocks [1, 2]. Plant viruses can be easily multiplied, which are structurally uniform, robust, and biodegradable with a size particularly suitable for nanoscale applications. Virus-like particles (VLPs) are multimeric self-assembled protein complexes mimicking the organization and conformation of native viruses but lack the viral genome making them replication-deficient and noninfectious [3]. VLPs consist of protein shells (termed as capsids), and the capsids are typically composed of identical coat protein subunits. Peptide-based vaccines are in general poorly immunogenic and for this reason they require multiple injections and adjuvants in order to increase their effectiveness. VLPs offer a promising approach to the production of vaccines against many diseases, because their repetitive, high density display of epitopes is potentially highly effective in eliciting strong immune responses [4]. VLPs lacking viral nucleic acid are noninfectious. Nevertheless, they are self-assembled protein structures mimicking infectious viruses and thus constitute a safe and effective approach for the induction of neutralizing antibodies to surface proteins, where soluble forms of their protein subunits have failed. It has been also reported that viral structures are regarded as a vaccine platform to display foreign epitopes [5].

In general, bacteria, yeast, insect, and animal cells have been applied as cell-based systems to produce VLPs. The bacterial cell cultures have been explored as a VLP production platform with advantages in terms of scalability and production cost [6]. However, bacteria are prokaryotes which lack glycosylation process essential for proper immunogenicity and antigen stability when VLPs are applied as vaccines. In contrast to bacteria, yeast cells have glycosylation apparatus [7]. However, their glycoforms are mainly high mannose type, which is not desirable for the most therapeutic glycoproteins [8]. The matured glycoforms in baculovirus-insect cell system also are mainly high mannose type [9]. The glycosylation of envelope proteins affects their folding and thus is essential for formation and immunogenicity of VLPs [10, 11]. In glycosylation process, bacteria, yeast, and insect cells have fundamental limitations. The mammalian cells have proper glycosylation apparatus and ability to fold the envelope proteins of virus, which facilitate functional VLPs production. However, the mammalian cell-based systems require manufacturing facilities including fermentation bioreactors for large-scale upstream processing, which is too expensive to establish. This high production cost is a major disadvantage of the mammalian cell-based system. Plants do not need such expensive facilities to produce biomass. Thus, plants are considered as a potential bioreactor system for VLPs with advantages such as low cost of upstream biomass process, flexible production scalability, and the lack of human pathogen contaminants [12, 13]. Nevertheless, plants for VLP production platform are not perfectly acceptable due to relatively lower VLP production level than animal systems and plant-specific N-glycosylation of glycoproteins [14, 15]. Development of new plant expression system and advanced N-glycosylation engineering overcome such hurdles.

2. Virus-Like Particles in Plant Expression Systems

VLPs can be generated through different types of viral vectors and expression strategies in plants [16, 17]. The plant-derived viral vectors used for VLP expression can be classified into full virus vectors such as the potato virus X (PVX) [18, 19] and the cowpea mosaic virus (CPMV) [20] and the deconstructed vectors such as bean yellow dwarf virus (BeYDV) [21, 22] and MagniCON based on tobacco mosaic virus (TMV) [23, 24]. The earlier plant VLPs were Hepatitis B core antigen (HBcAg) VLPs [25] and Hepatitis B surface antigen (HBsAg) VLPs fused to soybean vegetable storage protein vspA (VSPαS) in transgenic tobacco leaves obtained by Agrobacterium-mediated transformation [25, 26]. PVX and CPMV based viral vectors were applied to generate HBcAg VLPs [27]. Transgenic plants using Agrobacterium-mediated DNA transfer have been used for the stable gene expression system for VLPs; however the VLP expression level is low [10~24 μg/g fresh leaf weight (FLW)] [27, 28]. The human papilloma virus (HPV) L-1 based VLP vaccines were also successfully expressed and assembled in stable transgenic potato and tobacco [2931]. Unlike stable transformation, the transient expression with deconstructed geminiviral vectors showed 80 times higher accumulation of HBcAg VLPs in N. benthamiana compared to both PVX and CPMV vectors [32]. Another transient expression system using deconstructed TMV-based MagnICON vector agroinfiltration has allowed the production of HBcAg VLPs with a yield of 2.38 mg/g (FLW), almost 3 times higher compared to the geminiviral vector within a short period time [17, 3235]. The intraperitoneally injected HBcAg VLPs obtained from the MagnICON system efficiently induced immune responses generating HBcAg specific IgGs in mice. These results indicate that, among currently available VLP expression systems, the production of large quantities of VLPs for vaccine applications is more feasible using MagniCON systems. In many cases genetic manipulation of plant-derived VLPs has been performed to modify the external surface of the particle. To this end, the heterologous polypeptide has been fused at the N- or C-terminus of the CP. VLPs can also be exploited as “platforms” for the presentation of foreign epitopes and/or targeting molecules on chimeric VLPs (cVLPs) [1, 2, 6]. Indeed, the VLPs can display multicomponent vaccine candidate epitopes as a fusion form between two different proteins [5]. For instance, the green fluorescent protein (GFP) and the HB surface antigen (HBsAg) S-protein were transiently expressed and heterodimerized with the native HBsAg sequentially forming chimeric VLPs (cVLPs) in N. benthamiana [36]. The HBsAg fusion with GFP was showed to be more stable and immunogenic than native HBsAg in in vivo mice experiment, indicating that cVLPs can be applied to display heterologous antigens to generate more immunogenic vaccines [5]. The fusion proteins between domain III (DIII) of West Nile virus (WNV) and HBcAg were expressed and displayed as cVLPs with geminiviral transient expression vectors in N. benthamiana [37]. In addition, the influenza virus M2 epitope [38] or HPV16 epitopes [27, 39, 40] individually were fused to HBcAg induced strong immune responses generating specific antibodies. The cVLPs displaying both HPV16 E6 and E7 proteins triggered their specific antibodies, respectively [39]. In general, vaccines are administered through intramuscular, subcutaneous, and intravenous injections. In addition, vaccines can be orally or nasally applied to induce mucosal immune responses [17]. Indeed, various results indicate that VLPs can be applied safely as oral vaccines carrying multiple epitopes without needle injection. For example, oral delivery of purified Norwalk virus CP (NVCP) VLPs produced in tobacco and tomato stimulated mucosal and serum immune responses to produce IgA and IgG [41] and oral administration with HBsAg displaying HIV-1 ENV and GAG epitopes provoked strong serum and mucosal antibody responses in mice [42]. These results indicate that VLPs can be applied safely as oral vaccines carrying multiple epitopes without needle injection.

3. Glycosylation of VLP Vaccines

Even though virus-like particles- (VLPs-) based vaccines have shown promising results, commercial production systems are currently limited to eukaryotic cells such as yeast, insect, and mammalian [14]. For instance, Lassa virus (LASV) VLPs cannot be easily produced in bacterial cell systems, because bacteria are incapable of performing glycosylation and other posttranslational protein modifications which are a key feature in most VLP-based proteins [14]. The glycosylation pattern of GP1 and GP2 glycoproteins of Lassa virus (LASV) has been shown to play a critical structural and functional role in preserving protein stability and allowing binding and fusion to host cells [43]. The glycosylation of VLP proteins has major impact on their structure and function, and thus it is important to determine the choice of platforms for their production. As the viral glycoproteins localize, guide, and potentiate the process of enveloped virus assembly, it becomes important to study their individual and combined behavior upon expression in both animal and plant cells, in order to identify domains within the glycoproteins responsible for the critical differences between the intracellular targeting in either cell system. The large structural protein of lettuce necrotic yellow virus was glycosylated with complex oligosaccharides containing N-acetylglucosamine N-linked to asparagine residues [44]. The potato virus X CP and PPV CP were also glycosylated [45]. Glycosylated CP of beet western yellows virus plays a role in the virus/aphis interaction and promotes the aphid transmission of the virus [46]. Reviewed earlier, plants offer an attractive alternative system for VLP vaccine production with cost-effective, scalable, versatile, appropriate glycosylation, efficient assembly of VLP, and safety from adventitious human pathogens [12].

Although it yielded encouraging results, expression of VLPs expressed in plants suffers from plant-specific glycosylation of glycoproteins [14, 15]. Most proteins in eukaryotic multicellular organisms including plants are synthesized as glycoproteins with N- and O-glycosylation, which are important posttranslational protein modifications [47]. N-glycans attached to proteins are crucial for protein folding, assembly, and their stability but also involved in cell to cell adhesion, protein targeting, and immune responses as biological activity [48, 49]. During the glycoprotein transportation through the secretory pathway, the oligosaccharide N-linked to the asparagine residue (Asn) undergoes several maturation steps involving the removal of glucose and mannose residues by different exoglycosidase to generate high mannose type N-glycan in the endoplasmic reticulum (ER) and the Golgi apparatus and, eventually, it is characterized by the addition of new oligosaccharide residues in the Golgi apparatus to form the matured complex type N-glycan (Figure 1). In the ER, the first step of N-glycosylation of plant proteins is the transfer of the oligosaccharide precursor Glc3Man9GlcNAc2 from a dolichol lipid to specific Asn residues on the nascent polypeptide chain [50]. Processing of this oligosaccharide into high mannose, complex, hybrid, or paucimannosidic type N-glycan occurs during the secretory pathway. Particularly in plant, the β-mannose is substituted by a bisecting β1,2-xylose that is not found in mammalian N-glycans, and the proximal N-acetylglucosamine of the core is substituted by an α1,3-fucose, instead of an α1,6-fucose in mammalians. In addition, β1,3-galactose and fucose that are α1,4-linked to the terminal N-acetylglucosamine of plant N-glycans form Lewis a () glycosylation (Figure 1) [51]. These modifications are not present in mammalian. Many mammalian complex N-glycans have an α1,6-fucose on the first core N-acetylglucosamine of N-glycan and are characterized by terminal β1,4-galactose and sialic acid which are not observed in plants (Figure 1) [5254]. Most plant-derived therapeutic proteins are complex glycoproteins requiring posttranslational modifications. The β1,2-xylose, core α1,3-fucose, and containing epitopes have been considered as immunogenic glycan epitopes found in plant-specific N-glycans. Such glycan residues are not present in humans, and thus proteins could cause immune rejection inducing plant-glycan specific antibodies causing protein clearance in blood stream as well as potential allergenic effects [5456]. These hurdles can be overcome by recent progress in plant glycoengineering. The plant expression with glycoengineering will allow the novel application of plant-made VLPs, including vessels for the delivery of small therapeutics, DNA fragments, and adjuvants (Figure 2).

4. N-Glycomodification in Plants

4.1. Targeted Expression to the ER

N-glycan structures influence biofunctionality and stability of therapeutic proteins and even directly affect immunogenicity of glycosylated subunit vaccines displayed on VLP surfaces. In plants, thus, N-glycosylation pathway has been modified in order to humanize the glycan structures of glycoproteins [57, 58].

A commonly used approach to express recombinant glycoproteins in plants is their accumulation in ER by addition of C-terminal signal H/KDEL ER retention motif [59]. The ER-retained proteins contain high mannose type N-glycans structurally similar between plant and mammalian cells [58, 60, 61]. The high mannose type N-glycans are oligosaccharide structures that mammals and plants have in common and thus are probably not immunogenic [62]. This strategy is largely devoid of plant-specific, immunogenic β1,2-xylose and core α1,3-fucose. Additionally, some studies have reported enhanced accumulation of KDEL-tagged proteins in the ER. Such ER retention of proteins usually increases the production level compared to that without KDEL in plant [63, 64]. Plant-derived monoclonal antibody (mAb) with high mannose N-glycan structure has shorter half-life than that of the mammalian-derived mAb with mammalian specific glycan structures [65]. However, mAb with high mannose glycans had relatively similar biological activities compared to the mammalian-derived mAb overcoming concerns about plant-specific glycoepitopes expressed by others [58]. In addition, the high mannose type glycan structure would be expected to cause an enhanced immune response through the mannose receptor (MR) on macrophages and dendritic cells recognizing the oligomannose of glycoproteins [66], which is an advantage for vaccine development. According to a previous study [67], the high mannose glycans on antigenic protein can render the protein more immunogenic, producing IgG against the high mannose glycosylated protein.

4.2. Knockout of Plant-Specific Glycosyltransferases

Gene inactivation or silencing may be used to reduce or eliminate the activity of plant- specific glycosyltransferases. In a plant cell, the specific enzymes are β1,2-xylosyltransferase and core α1,3-fucosyltransferase, which are responsible for transfer of the plant-specific xylose and fucose onto the attached N-glycan. Such glycan residues are not present in humans and are thus unwanted on proteins intended for therapeutic use. The knockout of the genes that are responsible for the synthesis of these glycan epitopes β1,2-xylosyltransferase and core α1,3-fucosyltransferase provides an easy strategy to solve this problem. The feasibility of this strategy has been proven by the generation of knockout Arabidopsis thaliana plant lacking xylosyltransferase and fucosyltransferase [68, 69]. In addition, biological activity assays of such glycoengineered mAbs showed that their antigen binding activity was not altered but significantly enhanced antibody-dependent cell-mediated cytotoxicity (ADCC) effect [70, 71]. Therapeutic antibodies without fucosylation have higher binding affinity for FcγRIIIa than for fucosylated human serum IgG, which is desirable to overcome the interference by human plasma IgG. Thus, the therapeutic antibodies without fucosylation can avoid the inhibitory effect of human plasma IgG on ADCC through their high FcγRIIIa binding affinity.

4.3. Humanization of Plant N-Glycosylation

The immunogenic and allergenic reactions of the β1,2-xylose and core α1,3-fucose N-glycan epitopes on plant-derived glycoproteins have been a problem for application of therapeutic proteins produced from plant expression system [72]. Glycoengineering strategies using transgenic plants and the availability of mutant plants lacking xylosyltransferase and fucosyltransferase genes for humanization of N-glycosylation allow producing recombinant proteins with more mammalian-like N-glycan structures in plant expression system. Most proteins used for therapy of human diseases are glycosylated, and the glycan structures have been shown to affect safety and efficacy of therapeutic glycoproteins [73]. Particularly, nonsialylation significantly causes shorter in vivo half-life of circulating glycoproteins, because exposed galactose glycan residues are recognized and captured by asialoglycoprotein receptors resulting in internalization of the glycoproteins in hepatocytes [74]. Terminal acid residues in N-linked glycans of most therapeutic glycoproteins affect important roles in in vivo physical stability, immunogenicity, and enzymatic activity [68, 70, 71, 74, 75]. Previous studies have demonstrated the importance of fully sialylated N-linked glycans and of consistency of homogeneous N-linked glycan structures on therapeutic glycoproteins in heterologous expression systems [7577]. For instance, the sialylated recombinant erythropoietin (EPO) had longer plasma half-life (5-6 h) compared to that (2 min) of desialylated EPO [78]. This beneficial effect of sialic acid on protein stability likely explains why knockin strategies for plant glycoengineering in glycosylation have mainly focused on the addition of terminal β1,4-galactose and sialic acid residues to humanize N-glycan in mutant plants lacking plant-specific N-glycan residues [75]. It was claimed that plant virus-based transient expression systems can be applied as the knockin strategy of β1,4-galactose and sialic acid transferring genes in the mutant plants, allowing the generation of abundant amount of therapeutic proteins within 1 week after virus infection, provide a feasible advantage over existing glycoprotein expression systems [75].

5. Conclusions

Taken together, plant-derived VLPs are considered safe because plants do not bear human pathogens and promising in terms of cost-effective scalability and speed of production. In fact, as far as upstream and downstream processing are concerned, plant-derived VLPs can take advantage of what has been done so far in the broader field of plant-made pharmaceuticals. Also compared to prokaryotes host cells, plants host guarantees the appropriate posttranslational modifications, such as glycosylation, often needed for proper protein function.

In plants, glycoengineering has been improved to create plants able to perform the ideal glycosylation enhancing efficacy and potency of VLPs-based therapeutics. As described in this review, several strategies, focused on the inactivation and/or addition of key enzymes, can be adopted to decorate tailor-made glycoforms of VLPs in plants. Thus, plant expression systems will be further improved for production of VLPs-based vaccines with respect to their proper glycomodification and the rapid and cost-effective expression.

Conflict of Interests

The authors declare that there is no conflict of interests regarding the publication of this paper.


This research was supported by Evaluation for Technology of Food, Agriculture, Forestry, and Fisheries, Korea (iPET-Code nos. 312037-5 and 111096-03-1-SB010) and a Grant (Code no. PJ009419022014) from the Korean Rural Development Administration, Korea institute of Planning.


  1. C. Lico, A. Schoubben, S. Baschieri, P. Blasi, and L. Santi, “Nanoparticles in biomedicine: new insights from plant viruses,” Current Medicinal Chemistry, vol. 20, no. 28, pp. 3471–3487, 2013. View at: Publisher Site | Google Scholar
  2. S. Grasso and L. Santi, “Viral nanoparticles as macromolecular devices for new therapeutic and pharmaceutical approaches,” International Journal of Physiology, Pathophysiology and Pharmacology, vol. 2, no. 2, pp. 161–178, 2010. View at: Google Scholar
  3. J. K. Pokorski and N. F. Steinmetz, “The art of engineering viral nanoparticles,” Molecular Pharmaceutics, vol. 8, no. 1, pp. 29–43, 2011. View at: Publisher Site | Google Scholar
  4. A. S. Bachmann, G. Corpuz, W. P. Hareld, G. Wang, and B. A. Coller, “A simple method for the rapid purification of copia virus-like particles from Drosophila Schneider 2 cells,” Journal of Virological Methods, vol. 115, no. 2, pp. 159–165, 2004. View at: Publisher Site | Google Scholar
  5. Q. Chen and H. Lai, “Plant-derived virus-like particles as vaccines,” Human Vaccines and Immunotherapeutics, vol. 9, no. 1, pp. 26–49, 2013. View at: Publisher Site | Google Scholar
  6. N. Kushnir, S. J. Streatfield, and V. Yusibov, “Virus-like particles as a highly efficient vaccine platform: diversity of targets and production systems and advances in clinical development,” Vaccine, vol. 31, no. 1, pp. 58–83, 2012. View at: Publisher Site | Google Scholar
  7. A. Herscovics and P. Orlean, “Glycoprotein biosynthesis in yeast,” The FASEB Journal, vol. 7, no. 6, pp. 540–550, 1993. View at: Google Scholar
  8. S. Wildt and T. U. Gerngross, “The humanization of N-glycosylation pathways in yeast,” Nature Reviews Microbiology, vol. 3, no. 2, pp. 119–128, 2005. View at: Publisher Site | Google Scholar
  9. R. L. Harrison and D. L. Jarvis, “Protein N-glycosylation in the baculovirus-insect cell expression system and engineering of insect cells to produce “mammalianized” recombinant glycoproteins,” Advances in Virus Research, vol. 68, pp. 159–191, 2006. View at: Publisher Site | Google Scholar
  10. J. Schwarzer, E. Rapp, and U. Reichl, “N-glycan analysis by CGE-LIF: profiling influenza A virus hemagglutinin N-glycosylation during vaccine production,” Electrophoresis, vol. 29, no. 20, pp. 4203–4214, 2008. View at: Publisher Site | Google Scholar
  11. S. E. Hamby and J. D. Hirst, “Prediction of glycosylation sites using random forests,” BMC Bioinformatics, vol. 9, article 500, 2008. View at: Publisher Site | Google Scholar
  12. L. Faye and V. Gomord, “Success stories in molecular farming-a brief overview,” Plant Biotechnology Journal, vol. 8, no. 5, pp. 525–528, 2010. View at: Publisher Site | Google Scholar
  13. G. Giddings, G. Allison, D. Brooks, and A. Carter, “Transgenic plants as factories for biopharmaceuticals,” Nature Biotechnology, vol. 18, no. 11, pp. 1151–1155, 2000. View at: Publisher Site | Google Scholar
  14. A. Roldao, M. C. Mellado, L. R. Castilho, M. J. Carrondo, and P. M. Alves, “Virus-like particles in vaccine development,” Expert Review of Vaccines, vol. 9, no. 10, pp. 1149–1176, 2010. View at: Publisher Site | Google Scholar
  15. H. Lai and Q. Chen, “Bioprocessing of plant-derived virus-like particles of Norwalk virus capsid protein under current Good Manufacture Practice regulations,” Plant Cell Reports, vol. 31, no. 3, pp. 573–584, 2012. View at: Publisher Site | Google Scholar
  16. Y. Gleba, V. Klimyuk, and S. Marillonnet, “Viral vectors for the expression of proteins in plants,” Current Opinion in Biotechnology, vol. 18, no. 2, pp. 134–141, 2007. View at: Publisher Site | Google Scholar
  17. L. Santi, L. Batchelor, Z. Huang et al., “An efficient plant viral expression system generating orally immunogenic Norwalk virus-like particles,” Vaccine, vol. 26, no. 15, pp. 1846–1854, 2008. View at: Publisher Site | Google Scholar
  18. S. Chapman, T. Kavanagh, and D. Baulcombe, “Potato virus X as a vector for gene expression in plants,” The Plant Journal, vol. 2, no. 4, pp. 549–557, 1992. View at: Google Scholar
  19. N. Cerovska, H. Hoffmeisterova, T. Moravec et al., “Transient expression of Human papillomavirus type 16 L2 epitope fused to N- and C-terminus of coat protein of Potato virus X in plants,” Journal of Biosciences, vol. 37, no. 1, pp. 125–133, 2012. View at: Publisher Site | Google Scholar
  20. A. Chatterji, W. F. Ochoa, M. Paine, B. R. Ratna, J. E. Johnson, and T. Lin, “New addresses on an addressable virus nanoblock: uniquely reactive Lys residues on cowpea mosaic virus,” Chemistry and Biology, vol. 11, no. 6, pp. 855–863, 2004. View at: Publisher Site | Google Scholar
  21. X. Zhang and H. Mason, “Bean Yellow Dwarf Virus replicons for high-level transgene expression in transgenic plants and cell cultures,” Biotechnology and Bioengineering, vol. 93, no. 2, pp. 271–279, 2006. View at: Publisher Site | Google Scholar
  22. L. Liu, J. W. Davies, and J. Stanley, “Mutational analysis of bean yellow dwarf virus, a geminivirus of the genus Mastrevirus that is adapted to dicotyledonous plants,” Journal of General Virology, vol. 79, pp. 2265–2274, 1998. View at: Google Scholar
  23. M. L. Smith, J. A. Lindbo, S. Dillard-Telm et al., “Modified tobacco mosaic virus particles as scaffolds for display of protein antigens for vaccine applications,” Virology, vol. 348, no. 2, pp. 475–488, 2006. View at: Publisher Site | Google Scholar
  24. G. Conti, M. C. Rodriguez, C. A. Manacorda, and S. Asurmendi, “Transgenic expression of Tobacco mosaic virus capsid and movement proteins modulate plant basal defense and biotic stress responses in Nicotiana tabacum,” Molecular Plant-Microbe Interactions, vol. 25, no. 10, pp. 1370–1384, 2012. View at: Publisher Site | Google Scholar
  25. S. Tsuda, K. Yoshioka, T. Tanaka et al., “Application of the human hepatitis B virus core antigen from transgenic tobacco plants for serological diagnosis,” Vox Sanguinis, vol. 74, no. 3, pp. 148–155, 1998. View at: Publisher Site | Google Scholar
  26. P. Sojikul, N. Buehner, and H. S. Mason, “A plant signal peptide-hepatitis B surface antigen fusion protein with enhanced stability and immunogenicity expressed in plant cells,” Proceedings of the National Academy of Sciences of the United States of America, vol. 100, no. 5, pp. 2209–2214, 2003. View at: Publisher Site | Google Scholar
  27. I. A. Mechtcheriakova, M. A. Eldarov, L. Nicholson, M. Shanks, K. G. Skryabin, and G. P. Lomonossoff, “The use of viral vectors to produce hepatitis B virus core particles in plants,” Journal of Virological Methods, vol. 131, no. 1, pp. 10–15, 2006. View at: Publisher Site | Google Scholar
  28. Z. Huang and H. S. Mason, “Conformational analysis of hepatitis B surface antigen fusions in an Agrobacterium-mediated transient expression system,” Plant Biotechnology Journal, vol. 2, no. 3, pp. 241–249, 2004. View at: Publisher Site | Google Scholar
  29. H. Warzecha, H. S. Mason, C. Lane et al., “Oral immunogenicity of human papillomavirus-like particles expressed in potato,” Journal of Virology, vol. 77, no. 16, pp. 8702–8711, 2003. View at: Publisher Site | Google Scholar
  30. A. Varsani, A. L. Williamson, D. Stewart, and E. P. Rybicki, “Transient expression of Human papillomavirus type 16 L1 protein in Nicotiana benthamiana using an infectious tobamovirus vector,” Virus Research, vol. 120, no. 1-2, pp. 91–96, 2006. View at: Publisher Site | Google Scholar
  31. A. Varsani, A.-L. Williamson, R. C. Rose, M. Jaffer, and E. P. Rybicki, “Expression of Human papillomavirus type 16 major capsid protein in transgenic Nicotiana tabacum cv. Xanthi,” Archives of Virology, vol. 148, no. 9, pp. 1771–1786, 2003. View at: Publisher Site | Google Scholar
  32. Z. Huang, Q. Chen, B. Hjelm, C. Arntzen, and H. Mason, “A DNA replicon system for rapid high-level production of virus-like particles in plants,” Biotechnology and Bioengineering, vol. 103, no. 4, pp. 706–714, 2009. View at: Publisher Site | Google Scholar
  33. J. He, H. Lai, C. Brock, and Q. Chen, “A novel system for rapid and cost-effective production of detection and diagnostic reagents of West Nile virus in plants,” Journal of Biomedicine and Biotechnology, vol. 2012, Article ID 106783, 10 pages, 2012. View at: Publisher Site | Google Scholar
  34. K. L. Hefferon, “Broadly neutralizing antibodies and the promise of universal vaccines: where are we now?” Immunotherapy, vol. 6, no. 1, pp. 51–57, 2014. View at: Publisher Site | Google Scholar
  35. Z. Huang, L. Santi, K. LePore, J. Kilbourne, C. J. Arntzen, and H. S. Mason, “Rapid, high-level production of hepatitis B core antigen in plant leaf and its immunogenicity in mice,” Vaccine, vol. 24, no. 14, pp. 2506–2513, 2006. View at: Publisher Site | Google Scholar
  36. C. Lambert, N. Thome, C. J. Kluck, and R. Prange, “Functional incorporation of green fluorescent protein into hepatitis B virus envelope particles,” Virology, vol. 330, no. 1, pp. 158–167, 2004. View at: Publisher Site | Google Scholar
  37. Q. Chen, J. He, W. Phoolcharoen, and H. S. Mason, “Geminiviral vectors based on bean yellow dwarf virus for production of vaccine antigens and monoclonal antibodies in plants,” Human Vaccines, vol. 7, no. 3, pp. 331–338, 2011. View at: Publisher Site | Google Scholar
  38. N. V. Ravin, R. Y. Kotlyarov, E. S. Mardanova et al., “Plant-produced recombinant influenza vaccine based on virus-like HBc particles carrying an extracellular domain of M2 protein,” Biochemistry, vol. 77, no. 1, pp. 33–40, 2012. View at: Publisher Site | Google Scholar
  39. G. P. de la Rosa, A. Monroy-Garcia, M. de Lourdes Mora-García et al., “An HPV 16 L1-based chimeric human papilloma virus-like particles containing a string of epitopes produced in plants is able to elicit humoral and cytotoxic T-cell activity in mice,” Virology Journal, vol. 6, article 2, 2009. View at: Publisher Site | Google Scholar
  40. L. Santi, Z. Huang, and H. Mason, “Virus-like particles production in green plants,” Methods, vol. 40, no. 1, pp. 66–76, 2006. View at: Publisher Site | Google Scholar
  41. Z. Huang, G. Elkin, B. J. Maloney et al., “Virus-like particle expression and assembly in plants: hepatitis B and Norwalk viruses,” Vaccine, vol. 23, no. 15, pp. 1851–1858, 2005. View at: Publisher Site | Google Scholar
  42. S. N. Shchelkunov, R. K. Salyaev, S. G. Pozdnyakov et al., “Immunogenicity of a novel, bivalent, plant-based oral vaccine against hepatitis B and human immunodeficiency viruses,” Biotechnology Letters, vol. 28, no. 13, pp. 959–967, 2006. View at: Publisher Site | Google Scholar
  43. L. M. Branco, J. N. Grove, F. J. Geske et al., “Lassa virus-like particles displaying all major immunological determinants as a vaccine candidate for Lassa hemorrhagic fever,” Virology Journal, vol. 7, article 279, 2010. View at: Publisher Site | Google Scholar
  44. R. G. Dietzgen and R. I. Francki, “Analysis of lettuce necrotic yellows virus structural proteins with monoclonal antibodies and concanavalin A,” Virology, vol. 166, no. 2, pp. 486–494, 1988. View at: Google Scholar
  45. L. A. Baratova, N. V. Fedorova, E. N. Dobrov et al., “N-terminal segment of potato virus X coat protein subunits is glycosylated and mediates formation of a bound water shell on the virion surface,” European Journal of Biochemistry, vol. 271, no. 15, pp. 3136–3145, 2004. View at: Publisher Site | Google Scholar
  46. P. Seddas and S. Boissinot, “Glycosylation of beet Western yellows virus proteins is implicated in the aphid transmission of the virus,” Archives of Virology, vol. 151, no. 5, pp. 967–984, 2006. View at: Publisher Site | Google Scholar
  47. A. Helenius and M. Aebi, “Intracellular functions of N-linked glycans,” Science, vol. 291, no. 5512, pp. 2364–2369, 2001. View at: Publisher Site | Google Scholar
  48. P. Lerouge, M. Bardor, S. Pagny, V. Gomord, and L. Faye, “N-glycosylation of recombinant pharmaceutical glycoproteins produced in transgenic plants: towards an humanisation of plant N-glycans,” Current Pharmaceutical Biotechnology, vol. 1, no. 4, pp. 347–354, 2000. View at: Google Scholar
  49. A. Varki, “Biological roles of oligosaccharides: all of the theories are correct,” Glycobiology, vol. 3, no. 2, pp. 97–130, 1993. View at: Google Scholar
  50. V. Gomord, A. Fitchette, L. Menu-Bouaouiche et al., “Plant-specific glycosylation patterns in the context of therapeutic protein production,” Plant Biotechnology Journal, vol. 8, no. 5, pp. 564–587, 2010. View at: Publisher Site | Google Scholar
  51. C. Saint-Jore-Dupas, L. Faye, and V. Gomord, “From planta to pharma with glycosylation in the toolbox,” Trends in Biotechnology, vol. 25, no. 7, pp. 317–323, 2007. View at: Publisher Site | Google Scholar
  52. M. Bencurova, W. Hemmer, M. Focke-Tejkl, I. B. Wilson, and F. Altmann, “Specificity of IgG and IgE antibodies against plant and insect glycoprotein glycans determined with artificial glycoforms of human transferrin,” Glycobiology, vol. 14, no. 5, pp. 457–466, 2004. View at: Publisher Site | Google Scholar
  53. K. Fotisch and S. Vieths, “N- and O-linked oligosaccharides of allergenic glycoproteins,” Glycoconjugate Journal, vol. 18, no. 5, pp. 373–390, 2001. View at: Publisher Site | Google Scholar
  54. R. van Ree, M. Cabanes-Macheteau, J. Akkerdaas et al., “β(1,2)-xylose and α(1,3)-fucose residues have a strong contribution in IgE binding to plant glycoallergens,” The Journal of Biological Chemistry, vol. 275, no. 15, pp. 11451–11458, 2000. View at: Publisher Site | Google Scholar
  55. F. Altmann, “The role of protein glycosylation in allergy,” International Archives of Allergy and Immunology, vol. 142, no. 2, pp. 99–115, 2007. View at: Publisher Site | Google Scholar
  56. D. Bosch, A. Castilho, A. Loos, A. Schots, and H. Steinkellner, “N-glycosylation of plant-produced recombinant proteins,” Current Pharmaceutical Design, vol. 19, no. 31, pp. 5503–5512, 2013. View at: Publisher Site | Google Scholar
  57. R. Strasser, J. Stadlmann, B. Svoboda, F. Altmann, J. Glossl, and L. Mach, “Molecular basis of N-acetylglucosaminyltransferase I deficiency in Arabidopsis thaliana plants lacking complex N-glycans,” Biochemical Journal, vol. 387, pp. 385–391, 2005. View at: Publisher Site | Google Scholar
  58. K. Ko, Y. Tekoah, P. M. Rudd et al., “Function and glycosylation of plant-derived antiviral monoclonal antibody,” Proceedings of the National Academy of Sciences of the United States of America, vol. 100, no. 13, pp. 8013–8018, 2003. View at: Publisher Site | Google Scholar
  59. V. Gomord, L. A. Denmat, A. C. Fitchette-Laine, B. Satiat-Jeunemaitre, C. Hawes, and L. Faye, “The C-terminal HDEL sequence is sufficient for retention of secretory proteins in the endoplasmic reticulum (ER) but promotes vacuolar targeting of proteins that escape the ER,” The Plant Journal, vol. 11, no. 2, pp. 313–325, 1997. View at: Publisher Site | Google Scholar
  60. D. M. Floss, M. Sack, J. Stadlmann et al., “Biochemical and functional characterization of anti-HIV antibody-ELP fusion proteins from transgenic plants,” Plant Biotechnology Journal, vol. 6, no. 4, pp. 379–391, 2008. View at: Publisher Site | Google Scholar
  61. S. Petruccelli, M. S. Otegui, F. Lareu et al., “A KDEL-tagged monoclonal antibody is efficiently retained in the endoplasmic reticulum in leaves, but is both partially secreted and sorted to protein storage vacuoles in seeds,” Plant Biotechnology Journal, vol. 4, no. 5, pp. 511–527, 2006. View at: Publisher Site | Google Scholar
  62. V. Gomord, C. Sourrouille, A. C. Fitchette et al., “Production and glycosylation of plant-made pharmaceuticals: the antibodies as a challenge,” Plant Biotechnology Journal, vol. 2, no. 2, pp. 83–100, 2004. View at: Publisher Site | Google Scholar
  63. U. Conrad and U. Fiedler, “Compartment-specific accumulation of recombinant immunoglobulins in plant cells: an essential tool for antibody production and immunomodulation of physiological functions and pathogen activity,” Plant Molecular Biology, vol. 38, no. 1-2, pp. 101–109, 1998. View at: Google Scholar
  64. J. M. Sharp and P. M. Doran, “Strategies for enhancing monoclonal antibody accumulation in plant cell and organ cultures,” Biotechnology Progress, vol. 17, no. 6, pp. 979–992, 2001. View at: Publisher Site | Google Scholar
  65. J. H. Lee, D. Y. Park, K. J. Lee et al., “Intracellular reprogramming of expression, glycosylation, and function of a plant-derived antiviral therapeutic monoclonal antibody,” PLoS ONE, vol. 8, no. 8, Article ID e68772, 2013. View at: Publisher Site | Google Scholar
  66. V. Apostolopoulos and I. F. McKenzie, “Role of the mannose receptor in the immune response,” Current Molecular Medicine, vol. 1, no. 4, pp. 469–474, 2001. View at: Google Scholar
  67. Z. Lu, K. J. Lee, Y. Shao et al., “Expression of GA733-Fc fusion protein as a vaccine candidate for colorectal cancer in transgenic plants,” Journal of Biomedicine and Biotechnology, vol. 2012, Article ID 364240, 11 pages, 2012. View at: Publisher Site | Google Scholar
  68. M. Schahs, R. Strasser, J. Stadlmann, R. Kunert, T. Rademacher, and H. Steinkellner, “Production of a monoclonal antibody in plants with a humanized N-glycosylation pattern,” Plant Biotechnology Journal, vol. 5, no. 5, pp. 657–663, 2007. View at: Publisher Site | Google Scholar
  69. R. Strasser, F. Altmann, L. Mach, J. Glossl, and H. Steinkellner, “Generation of Arabidopsis thaliana plants with complex N-glycans lacking β1,2-linked xylose and core α1,3-linked fucose,” FEBS Letters, vol. 561, no. 1–3, pp. 132–136, 2004. View at: Publisher Site | Google Scholar
  70. K. M. Cox, J. D. Sterling, J. T. Regan et al., “Glycan optimization of a human monoclonal antibody in the aquatic plant Lemna minor,” Nature Biotechnology, vol. 24, no. 12, pp. 1591–1597, 2006. View at: Publisher Site | Google Scholar
  71. D. N. Forthal, J. S. Gach, G. Landucci et al., “Fc-glycosylation influences Fcγ receptor binding and cell-mediated anti-hivactivity of monoclonal antibody 2G12,” The Journal of Immunology, vol. 185, no. 11, pp. 6876–6882, 2010. View at: Publisher Site | Google Scholar
  72. M. Cabanes-Macheteau, A. Fitchette-Laine, C. Loutelier-Bourhis et al., “N-glycosylation of a mouse IgG expressed in transgenic tobacco plants,” Glycobiology, vol. 9, no. 4, pp. 365–372, 1999. View at: Google Scholar
  73. R. Apweiler, H. Hermjakob, and N. Sharon, “On the frequency of protein glycosylation, as deduced from analysis of the SWISS-PROT database,” Biochimica et Biophysica Acta, vol. 1473, no. 1, pp. 4–8, 1999. View at: Publisher Site | Google Scholar
  74. M. N. Fukuda, H. Sasaki, L. Lopez, and M. Fukuda, “Survival of recombinant erythropoietin in the circulation: the role of carbohydrates,” Blood, vol. 73, no. 1, pp. 84–89, 1989. View at: Google Scholar
  75. A. Castilho, R. Strasser, J. Stadlmann et al., “In planta protein sialylation through overexpression of the respective mammalian pathway,” The Journal of Biological Chemistry, vol. 285, no. 21, pp. 15923–15930, 2010. View at: Publisher Site | Google Scholar
  76. J. C. Egrie and J. K. Browne, “Development and characterization of novel erythropoiesis stimulating protein (NESP),” Nephrology Dialysis Transplantation, vol. 16, supplement 3, pp. 3–13, 2001. View at: Google Scholar
  77. F. A. Ngantung, P. G. Miller, F. R. Brushett, G. L. Tang, and D. I. Wang, “RNA interference of sialidase improves glycoprotein sialic acid content consistency,” Biotechnology and Bioengineering, vol. 95, no. 1, pp. 106–119, 2006. View at: Publisher Site | Google Scholar
  78. S. Erbayraktar, G. Grasso, A. Sfacteria et al., “Asialoerythropoietin is a nonerythropoietic cytokine with broad neuroprotective activity in vivo,” Proceedings of the National Academy of Sciences of the United States of America, vol. 100, no. 11, pp. 6741–6746, 2003. View at: Publisher Site | Google Scholar

Copyright © 2014 Hyun-Soon Kim et al. This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

More related articles

 PDF Download Citation Citation
 Download other formatsMore
 Order printed copiesOrder

Related articles

We are committed to sharing findings related to COVID-19 as quickly as possible. We will be providing unlimited waivers of publication charges for accepted research articles as well as case reports and case series related to COVID-19. Review articles are excluded from this waiver policy. Sign up here as a reviewer to help fast-track new submissions.