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Chloroplast-Derived Vaccine Antigens and Biopharmaceuticals: Expression, Folding, Assembly and Functionality

  • S. Chebolu
  • H. Daniell
Chapter
Part of the Current Topics in Microbiology and Immunology book series (CT MICROBIOLOGY, volume 332)

Chloroplast genetic engineering offers several advantages, including high levels of transgene expression, transgene containment via maternal inheritance, and multi-gene expression in a single transformation event. Oral delivery is facilitated by hyperexpression of vaccine antigens against cholera, tetanus, anthrax, plague, or canine parvovirus (4%–31% of total soluble protein, TSP) in transgenic chloroplasts (leaves) or non-green plastids (carrots, tomato) as well as the availability of antibiotic free selectable markers or the ability to excise selectable marker genes. Hyperexpression of several therapeutic proteins, including human serum albumin (11.1% TSP), somatotropin (7% TSP), interferon-alpha (19% TSP), interferon-gamma (6% TSP), and antimicrobial peptide (21.5% TSP), facilitates efficient and economic purification. Also, the presence of chap-erones and enzymes in chloroplasts facilitates assembly of complex multisubunit proteins and correct folding of human blood proteins with proper disulfide bonds. Functionality of chloroplast-derived vaccine antigens and therapeutic proteins has been demonstrated by several assays, including the macrophage lysis assay, GM1-ganglioside binding assay, protection of HeLA cells or human lung carcinoma cells against encephalomyocarditis virus, systemic immune response, protection against pathogen challenge, and growth or inhibition of cell cultures. Purification of human proinsulin has been achieved using novel purification strategies (inverse temperature transition property) that do not require expensive column chromatography techniques. Thus, transgenic chloroplasts are ideal bio-reactors for production of functional human and animal therapeutic proteins in an environmentally friendly manner.

Keywords

Human Serum Albumin Chloroplast Genome Total Soluble Protein Therapeutic Protein Oral Delivery 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.

References

  1. Arlen PA, Singleton M, Adamovicz JJ, Ding Y, Davoodi-Semiromi A, Daniell H (2008) Effective plague vaccination via oral delivery of plant cells expressing F1-V antigens in chloroplasts. Infect. Immun. 76:3640–3650PubMedCrossRefGoogle Scholar
  2. Arlen PA, Falconer R, Cherukumilli S, Cole A, Cole AM, Oishi KK, Daniell H (2007) Field production and functional evaluation of chloroplast-derived interferon-alpha2b. Plant Biotechnol. J. 5:511–525PubMedCrossRefGoogle Scholar
  3. Baillie L (2001) The development of new vaccines against Bacillus anthracis. J Appl Microbiol 91:609–613PubMedCrossRefGoogle Scholar
  4. Bogorad L (2000) Engineering chloroplasts: an alternative site for foreign genes, proteins, reactions and products. Trends Biotechnol 18:257–263PubMedCrossRefGoogle Scholar
  5. Daniell H (2002) Molecular strategies for gene containment in transgenic crops. Nat Biotechnol 20:581–586PubMedCrossRefGoogle Scholar
  6. Daniell H (2004) Medical Molecular Pharming. In: Goodman RM (ed) The encyclopedia of plant and crop sciences. Marcel Dekker, New York, pp 705–710Google Scholar
  7. Daniell H, Dhingra A (2002) Multigene engineering: dawn of an exciting new era in biotechnology. Curr Opin Biotechnol 13:136–141PubMedCrossRefGoogle Scholar
  8. Daniell H, Wycoff K (2001) WO Patent 01-64929, 2001Google Scholar
  9. Daniell H, Guda C, McPherson DT, Xu J, Zhang X, Urry DW (1997) Hyperexpression of an environmentally friendly synthetic polymer gene. Methods Mol Biol 63:359–371PubMedGoogle Scholar
  10. Daniell H, Dhingra A, Fernández-San Millán A (2001a) Chloroplast transgenic approach for production of antibodies, biopharmaceuticals and edible vaccines. In: 12th International Congress on Photosynthesis Vol. S40–04, CSIRO Publishing, Brisbane, Australia, pp 1–6Google Scholar
  11. Daniell H, Lee SB, Panchal T, Wiebe P (2001b) Expression of native cholera toxin B sub-unit gene and assembly as functional oligomers in transgenic tobacco chloroplasts. J Mol Biol 311:1001–1009CrossRefGoogle Scholar
  12. Daniell H, Muthukumar B, Lee SB (2001c) Marker free transgenic plants: engineering the chlo-roplast genome without the use of antibiotic selection. Curr Genet 39:109–116CrossRefGoogle Scholar
  13. Daniell H, Streatfield SJ, Wycoff K (2001d) Medical molecular farming: production of antibodies, biopharmaceuticals and edible vaccines in plants. Trends Plant Sci 6:219–226CrossRefGoogle Scholar
  14. Daniell H, Wiebe PO, Millan AF (2001e) Antibiotic-free chloroplast genetic engineering: an environmentally friendly approach. Trends Plant Sci 6:237–239CrossRefGoogle Scholar
  15. Daniell H (2004) Medical Molecular Pharming. In: Goodman RM (ed) The encyclopedia of plant and crop sciences. Marcel Dekker, New York, pp 705–710Google Scholar
  16. Daniell H, Carmona-Sanchez O, Burns B (2004a) Chloroplast derived antibodies, biopharmaceu-ticals and edible vaccines. In: Fischer R, Schillberg S (eds) Molecular farming. Wiley-VerlagVCH, Weinheim, Germany, pp 113–133CrossRefGoogle Scholar
  17. Daniell H, Watson J, Koya V, Leppla SH (2004b) Expression of Bacillus anthracis protective antigen in transgenic chloroplasts of tobacco, a non-food/feed crop. Vaccine 22:4374–4384CrossRefGoogle Scholar
  18. De Cosa B, Moar W, Lee SB, Miller M, Daniell H (2001) Over expression of the cry2Aa2 operon in chloroplasts leads to formation of insecticidal crystals. Nat Biotechnol 19:71–74PubMedCrossRefGoogle Scholar
  19. DeGray G, Rajasekaran K, Smith F, Sanford J, Daniell H (2001) Expression of an antimicrobial peptide via the chloroplast genome to control phytopathogenic bacteria and fungi. Plant Physiol 127:852–862PubMedCrossRefGoogle Scholar
  20. Dhingra A, Portis AR, Daniell H (2004) Enhanced translation of a chloroplast-expressed RbcS gene restores small subunit levels and photosynthesis in nuclear RbcS antisense plants. Proc Natl Acad Sci U S A 101:6315–6320PubMedCrossRefGoogle Scholar
  21. Elderbaum O, Stein D, Holland N, Gafni Y, Livneh O, Novick D, Rubinstein M, Sele I (1992) Expression of active human interferon beta in transgenic plants. J Interferon Res 12:449–453Google Scholar
  22. Falconer R (2002) Expression of Interferon alpha 2b in transgenic chloroplasts of a low-nicotine tobacco. M.S. thesis, University of Central FloridaGoogle Scholar
  23. Fernández-San Millán A, Mingo-Castel A, Miller M, Daniell H (2003) A chloroplast transgenic approach to hyper-express and purify Human Serum Albumin, a protein highly susceptible to proteolytic degradation. Plant Biotech J 1:71–79CrossRefGoogle Scholar
  24. Figueroa-Soto CG, Guillermo LC, Valenzuela-Soto EM (1999) Immunolocalization of betaine aldehyde dehydrogenase in porcine kidneys. Biochem Biophys Res Commun 258:732–736PubMedCrossRefGoogle Scholar
  25. Giddings G, Allison G, Brooks D, Carter A (2000) Transgenic plants as factories for biopharma-ceuticals. Nat Biotechnol 18:1151–1155PubMedCrossRefGoogle Scholar
  26. Gomez N, Carrillo C, Salinas J, Parra F, Borca M V, Escribano JM (1998) Expression of immuno-genic glycoprotein S polypeptides from transmissible gastroenteritis corona virus in transgenic plants. Virology 249:352–358PubMedCrossRefGoogle Scholar
  27. Guda C, Lee SB, Daniell H (2000) Stable expression of a protein based polymer in tobacco chlo-roplasts. Plant Cell Rep 19:257–262CrossRefGoogle Scholar
  28. Haq TA, Mason HS, Clements JD, Arntzen CJ (1995) Oral immunization with a recombinant bacterial antigen produced in transgenic plants. Science 268:714–716PubMedCrossRefGoogle Scholar
  29. Iatham S, Day A (2000) Removal of antibiotic resistance genes from transgenic tobacco plastids. Nat Biotechnol 18:1172–1176CrossRefGoogle Scholar
  30. Ivins B, Fellows P, Pitt L, Estep J, Farchaus J, Friedlander A, et al (1995) Experimental anthrax vaccines: efficacy of adjuvants combined with protective antigen against an aerosol Bacillus anthracis spore challenge in guinea pigs. Vaccine 13:1779–1783PubMedCrossRefGoogle Scholar
  31. Jacob L, Zasloff M (1994) Potential therapeutic applications of magainins and other microbial; agents animal origin: antimicrobial peptides. Ciba Found Symp 186:197–223PubMedGoogle Scholar
  32. Joellenback LM, Zwanziger LL, Durch JS, Strom BL (eds) (2002) “Anthrax vaccine manufacture” in the anthrax vaccine. Is it safe? Does it work? National Academy, Washington, DC, pp 180–197Google Scholar
  33. Kaufmann AF, Meltzer MI, Schmid GP (1997) The economic impact of a bioterrorist attack: are prevention and post attack intervention programs justifiable? Emerg Infect Dis 3:83–94PubMedCrossRefGoogle Scholar
  34. Klaus SM, Huang FC, Golds TJ, Koop HU (2004) Generation of marker-free plastid transformants using a transiently cointegrated selection gene. Nat Biotechnol 22:225PubMedCrossRefGoogle Scholar
  35. Kumar S, Dhingra A, Daniell H (2004) Plastid-expressed betaine aldehyde dehydrogenase gene in carrot cultured cells, roots, and leaves confers enhanced salt tolerance. Plant Physiol. 136:2843–2854.PubMedCrossRefGoogle Scholar
  36. Langeveld JP, Casal JI, Osterhaus AD, Cortes E, de Swart R, Vela C, Dalsgaard K, Puijk WC, Schaaper WM, Meloen RH (1994) First peptide vaccine providing protection against viral infection in the target animal: studies of canine parvovirus in dogs. J Virol 68:4506–4513PubMedGoogle Scholar
  37. Langeveld JP, Kamstrup S, Uttenthal A, Strandbygaard B, Vela C, Dalsgaard K, Beekman NJ, Meloen RH, Casal JI (1995) Full protection in mink against enteritis virus with new generation canine parvovirus vaccines based on synthetic peptide or recombinant protein. Vaccine 13:1033–1037PubMedCrossRefGoogle Scholar
  38. Larrick JW, Thomas DW (2001) Producing protein in transgenic plants and animals. Curr Opin Biotechnol 12:411–418PubMedCrossRefGoogle Scholar
  39. Lee SB, Kwon HB, Kwon SJ, Park SC, Jeong MJ, Han SE, Byun MO, Daniell H (2003) Accumulation of trehalose within transgenic chloroplasts confers drought tolerance. Mol Breed 11:1–13CrossRefGoogle Scholar
  40. Leelavathi S, Reddy VS (2003) Chloroplast expression of His-tagged GUS-fusions: a general strategy to overproduce and purify foreign proteins using transplastomic plants as bioreactors. Mol Breed 11:49–58CrossRefGoogle Scholar
  41. Mason HS, Lam D, Arntzen CJ (1992) Expression of hepatitis B surface antigen in transgenic plants. Proc Natl Acad Sci U S A 89:11745–11749PubMedCrossRefGoogle Scholar
  42. Mason HS, Haq TA, Clements JD, Arntzen CJ (1998) Edible vaccine protects mice against Escherichia coli heat-labile enterotoxin (LT): potatoes expressing a synthetic LT-B gene. Vaccine 16:1336–1343PubMedCrossRefGoogle Scholar
  43. Molina A, Hervas-Stubbs S, Daniell H, Mingo-Castel AM, Veramendi J (2004) High yield expression of a viral peptide animal vaccine in transgenic tobacco chloroplasts. Plant Biotechnol 2:141–153CrossRefGoogle Scholar
  44. Ruiz G (2002) Optimization of codon composition and regulatory elements for expression of the human IGF-1 in transgenic chloroplasts. MS thesis, University of Central FloridaGoogle Scholar
  45. Singleton ML (2003) Expression of CaF1 and LcrV as a fusion protein for a vaccine against Yersinia pestis via chloroplast genetic engineering. MS thesis, University of Central FloridaGoogle Scholar
  46. Tackaberry E, Dudani A, Prior F, Tocchi M, Sardana R, Altosaar I, Ganz PR (1999) Development of biopharmaceuticals in plant expression systems: cloning, expression and immunological reactivity of human cytomegalovirus glycoprotein B (UL55) in seeds of transgenic tobacco. Vaccine 17:3020–3029PubMedCrossRefGoogle Scholar
  47. Thanavala Y, Yang Y, Lyon P, Mason HS, Arntzen C (1995) Immunogenicity of transgenic plant-derived hepatitis B surface antigen. Proc Natl Acad Sci U S A 92:3358–3361PubMedCrossRefGoogle Scholar
  48. Titball RW, Williamson ED (2001) Vaccination against bubonic and pneumonic plague. Vaccine 19:4175–4184PubMedCrossRefGoogle Scholar
  49. Torrado J, Carrascosa C (2003) Pharmacological characteristics of parenteral IGF-I administration. Curr Pharm Biotechnol 4:123–140PubMedCrossRefGoogle Scholar
  50. Urry DW, McPherson DT, Xu J, Daniell H, Guda C, Gowda DC, Jing N, Parker TM (1996) Protein based polymeric materials: synthesis and properties. In: Salamone JC (ed) The polymeric materials encyclopedia: synthesis, properties and applications. CRC Press, Boca Raton, FL, pp 2645–2699Google Scholar
  51. Vivek BS, Ngo QA, Simon PW (1999) Evidence for maternal inheritance of the chloroplast genome in cultivated carrot (Daucus carota L. ssp. sativus). Theor Appl Genet 98:669–672CrossRefGoogle Scholar
  52. Williamson ED, Eley SM, Stagg AJ, Green M, Russell P, Titball RW (1997) A sub-unit vaccine elicits IgG in serum, spleen cell cultures and bronchial washings and protects immunized animals against plague. Vaccine 15:1079–1084PubMedCrossRefGoogle Scholar
  53. Zasloff M (1987) Magainins, a class of antimicrobial peptides from Xenopus skin: isolation, characterization of two active forms, and partial cDNA sequence of a precursor. Proc Natl Acad Sci U S A 84:5449–5953PubMedCrossRefGoogle Scholar
  54. Zhang Q, Liu Y, Sodmergen (2003) Examination of the cytoplasmic DNA in male reproductive cells to determine the potential for cytoplasmic inheritance in 295 angiosperm species. Plant Cell Physiol 44:941–951PubMedCrossRefGoogle Scholar

Copyright information

© Springer-Verlag Berlin Heidelberg 2009

Authors and Affiliations

  • S. Chebolu
    • 1
  • H. Daniell
    • 1
  1. 1.Department of Molecular Biology & MicrobiologyUniversity of Central FloridaOrlandoUSA

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