Transgenic Research

, Volume 9, Issue 1, pp 71–78 | Cite as

Expression of full-length bioactive antimicrobial human lactoferrin in potato plants

  • Daniel K.X. Chong
  • William H.R. Langridge


A cDNA fragment encoding human lactoferrin (hLF) linked to a plant microsomal retention signal peptide (SEKDEL) was stably integrated into the Solanum tuberosum genome by Agrobacterium tumefaciens-mediated leaf disk transformation methods. The lactoferrin gene was expressed under control of both the auxin-inducible manopine synthase (mas) P2 promoter and the cauliflower mosaic virus (CaMV) 35S tandem promoter. The presence of the hLF cDNA in the genome of regenerated transformed potato plants was detected by polymerase chain reaction amplification methods. Full-length hLF protein was identified by immunoblot analysis in tuber tissue extracts from the transformed plants by immunoblot analysis. The hLF produced in transgenic plant tissues migrated during polyacrylamide gel electrophoresis as a single band with an approximate molecular mass equal to hLF. Auxin activation of the mas P2 promoter increased lactoferrin expression levels in transformed tuber and leaf tissues to approximately 0.1% of total soluble plant protein. Antimicrobial activity against four different human pathogenic bacterial strains was detected in extracts of lactoferrin-containing potato tuber tissues. This is the first report of synthesis of full length, biologically active hLF in edible plants.

human lactoferrin potato transgenic plants 


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  1. Anderson BF, Baker HM, Norris GE, Rice DW and Baker EN (1989) Structure of human lactoferrin: crystallographic structure analysis and refinement at 2.8 Å resolution. J Mol Biol 209: 711–734.Google Scholar
  2. Arnold RR, Brewer M and Gauthier JJ (1980) Bacteriocidal activity of human lactoferrin: sensitivity of a variety of microorganisms. Infect Immun 28: 893–898.Google Scholar
  3. Arnold RR, Russell JE, Champion WJ and Gauthier JJ (1981) Bacteriocidal activity of human lactoferrin: influence of physical conditions and metabolic state of the target microorganism. Infect Immun 32: 655–660.Google Scholar
  4. Bellamy W, Takase M, Yamauchi K, Wakabayashi H, Kawase K and Tomita M (1992) Identification of the bacteriocidal domain of lactoferrin. Biochim Biophys Acta 1121: 130–136.Google Scholar
  5. Bennett RM and Davis J (1982) Lactoferrin interacts with deoxyribonucleic acid: a preferential reactivity with double-stranded DNA and dissociation of DNA-anti-DNA complexes. J Lab Clin Med 99: 127–138.Google Scholar
  6. Bradford MM (1976) A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem 72: 248–254.Google Scholar
  7. Broxmeyer HE, Mantel C, Gentile P, Srivastava C, Miyazawa K, Zucali JR, Rado TA, Levi S and Arosio P (1991) Actions of H-subunit ferritin and lactoferrin as suppressor molecules of myelopoiesis in vitro and in vivo. Curr Stud Hematol Blood Transfus 178–181.Google Scholar
  8. Chong DK, Roberts W, Arakawa T, Illes K, Bagi G, Slattery CW and Langridge WH (1997) Expression of the human milk protein beta-casein in transgenic potato plants. Transgenic Res 6: 289–296.Google Scholar
  9. Furmanski P, Li ZP, Fortuna MB, Swamy CV and Das MR (1989) Multiple molecular forms of human lactoferrin. Identification of a class of lactoferrins that possess ribonuclease activity and lack iron-binding capacity. J ExpMed 170: 415–429.Google Scholar
  10. Giugliano LG, Ribeiro ST, Vainstein MH and Ulhoa CJ (1995) Free secretory component and lactoferrin of human milk inhibit the adhesion of enterotoxigenic Escherichia coli. J Med Microbiol 42: 3–9.Google Scholar
  11. Hashizume S, Kuroda K and Murakami H (1983) Identification of lactoferrin as an essential growth factor for human lymphocytic cell lines in serum-free medium. Biochim Biophys Acta 763: 377–382.Google Scholar
  12. Kay R, Chen A, Daly M and McPherson J (1987) Duplication of CaMV 35S promoter sequences creates a strong enhancer for plant genes. Science 236: 1299–1302.Google Scholar
  13. Legrand D, Mazurier J, Metz-Boutigue MH, Jolles J, Jolles P, Montreuil J and Spik G (1984) Characterization and localization of an iron-binding 18-kDa glycopeptide isolated from the Nterminal half of human lactotransferrin. Biochim Biophys Acta 787: 90–96.Google Scholar
  14. Liang Q and Richardson (1993) Expression and characterization of human lactoferrin in yeast Saccharomyces cerevisiae. J Agric Food Chem 41: 1800–1807.Google Scholar
  15. Masson PL, Heremans JF and Ferin J (1968) Presence of an ironbinding protein (lactoferrin) in the genital tract of the human female. I. Its immunohistochemical localization in the endometrium. Fertil Steril 19: 679–689.Google Scholar
  16. Mitra A and Zhang Z (1994) Expression of a human lactoferrin cDNA in tobacco cells produces antibacterial protein(s). Plant Physiol 106: 977–981.Google Scholar
  17. Nuijens JH, van Berkel PH, Geerts ME, Hartevelt PP, de Boer HA, van Veen HA, and Pieper FR (1997) Characterization of recombinant human lactoferrin secreted in milk of transgenic mice. J Biol Chem 272: 8802–8807.Google Scholar
  18. Odell JT, Nagy F and Chua NH (1985) Identification of DNA sequences required for activity of the cauliflower mosaic virus 35S promoter. Nature 313: 810–812.Google Scholar
  19. Oseas R, Yang HH, Baehner RL and Boxer LA (1981) Lactoferrin: a promoter of polymorphonuclear leukocyte adhesiveness. Blood 57: 939–945.Google Scholar
  20. Platenburg GJ, Kootwijk EP, Kooiman PM, Woloshuk SL, Nuijens JH, Krimpenfort PJ, Pieper FR, de Boer HA and Strijker R (1994) Expression of human lactoferrin in milk of transgenic mice. Transgenic. Res. 3: 99–108.Google Scholar
  21. Saito H, Miyakawa H, Tamura Y, Shimamura S and Tomita M (1991) Potent bacteriocidal activity of bovine lactoferrin hydrolysate produced by heat treatment at acidic pH. J Dairy Sci 74: 3724–3730.Google Scholar
  22. Salmon V, Legrand D, Slomianny MC, el Yazidi I, Spik G, Gruber V, Bournat P, Olagnier B, Mison D, Theisen M and Merot B (1998) Production of human lactoferrin in transgenic tobacco plants. Protein Expr Purif 13: 127–135.Google Scholar
  23. Spik G, Strecker G, Fournet B, Bouquelet S, Montreuil J, Dorland L, van Halbeek H and Vliegenthart JF (1982) Primary structure of the glycans from human lactotransferrin. Eur J Biochem 121: 413–419.Google Scholar
  24. Stowell KM, Rado TA, Funk WD and Tweedie JW (1991) Expression of cloned human lactoferrin in baby-hamster kidney cells. Biochem J 276: 349–355.Google Scholar
  25. van Berkel PH, Geerts ME, van Veen HA, Kooiman PM, Pieper FR, de Boer HA and Nuijens JH (1995) Glycosylated and unglycosylated human lactoferrins both bind iron and show identical affinities towards human lysozyme and bacterial lipopolysaccharide, but differ in their susceptibilities towards tryptic proteolysis. Biochem J 312: 107–114.Google Scholar
  26. Velten J, Velten L and Schell J (1984) Isolation of a dual plant promoter fragment from the Ti plasmid of Agrobacterium tumefaciens. EMBO J 3: 2723–2730.Google Scholar
  27. Ward PP, May GS, Headon DR and Conneely OM (1992) An inducible expression system for the production of human lactoferrin in Aspergillus nidulans. Gene 122: 219–223.Google Scholar

Copyright information

© Kluwer Academic Publishers 2000

Authors and Affiliations

  • Daniel K.X. Chong
    • 1
  • William H.R. Langridge
    • 1
    • 2
  1. 1.Center for Molecular Biology and Gene TherapyLoma Linda
  2. 2.Department of BiochemistryLoma Linda UniversityLoma Linda

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