In silico investigation of lactoferrin protein characterizations for the prediction of anti-microbial properties

Document Type : Original article

Authors

1 Institute of Biotechnology Shiraz university

2 Head of Biotechnology Institute, Shiraz University, Shiraz, Iran

3 Institute of Biotechnology Shiraz University

4 Institute of Biotechnology Shiraz University School of Petroleum and Chemical engineering, Shiraz University, Shiraz, IR Iran

Abstract

Lactoferrin (Lf) is an iron-binding multi-functional glycoprotein which has numerous physiological functions such as iron transportation, anti-microbial activity and immune response. In this study, different in silico approaches were exploited to investigate Lf protein properties in a number of mammalian species. Results showed that the iron-binding site, DNA and RNA-binding sites, signal peptides and transferrin motifs in the Lf structure were highly conserved. Examined sequences showed three conserved motifs which were repeated twice in the Lf structure, demonstrating ancient duplication events in its gene. Also, results suggest that the functional domains in mammalian Lf proteins are Zinc finger, Tubulin/FtsZ, GTPase, α/β hydrolase and Zinc knuckle. The potential site for nucleic acid binding and the major DNA and RNA-binding sites in this protein were found in the lactoferricin (Lfc) fragment. Due to its high positive charge, Lf is able to bind a large number of compounds. Our analysis also revealed that the interactions between Lf and ITLN1, LYZ, CSN2, and CD14 proteins played an important role in the protective activities of Lf. Analysis for the prediction of secondary structures indicated that high amounts of α-helix, β-strand and β-sheet were present in Lf. The high degree of conservation among mammalian Lf proteins indicates that there is a close relationship between these proteins, reflecting their important role.

Keywords


1.Shanbacher FL, Goodman RE, Talhouk RS. Bovine mammary lactoferrin: Implications from messenger ribonucleic acid (mRNA) sequence and regulation contrary to other milk proteins. J Dairy Sci 1992; 76: 3812-3831.
2.Rodriguez DA, Vazquez L, Ramos G. Antimicrobial mechanisms and potential clinical application of lactoferrin [in Spanish]. Rev Latinoam Microbiol 2005; 47:102–11.
3.Wally J, Buchanan SK. A structural comparison of human serum transferrin and human lactoferrin. BioMetals 2007; 20: 249-262.
4.OztasYesim ER, Ozgunes N. Lactoferrin: a multifunctional protein. Adv Mol Med 2005; 1: 149-54.
5.Anderson BF, Baker HM, Dodson EJ, Norris GE, Rumball SV, Waters JM, Baker EN. Structure of human lactoferrin at 3.2-A° resolution. Proc Natl Acad Sci 1987; 84:1769-1773.
6.Khan JA, Kumar P, Paramasivam M, Yadav RS, Sahani MS, Sharma S, Srinivasana A, Singh TP. Camel lactoferrin, a transferrin-cum-lactoferrin: crystal structure of camel apolactoferrin at 2.6 Å resolution and structural basis of its dual role. J Mol Biol 2001; 309: 751-761.
7.Baker HM, Anderson BF, Baker EN. Dealing with iron: common structural principles in proteins that transport iron and heme. Proc Natl Acad Sci USA 2004; 100: 3579-3583.
8.Van der Strate BWA, Belijaars L, Molema G, Harmsen MC, Meijer DK. Antiviral activities of lactoferrin. Antiviral Res 2001; 52: 225–39.
9.Ellison RT, Giehl TJ, Laforce FM.  Damage of the membrane of enteric Gram-negative bacteria by lactoferrin and transferrin. Infect Immun 1988; 56: 2774-81.
10. Leitch EC, Willcox MDP. Elucidation of the antistaphylococcal action of lactoferrin and lysozyme. J med Microbial 1999; 48: 867-871.
11. Legrand D, Vigie K, Said EA, Elass E, Masson M, Slomianny MC. Surface nucleolin participates in both the binding and endocytosis of lactoferrin in target cells. Eur J Biochem 2004; 271: 303-17.
12. Bennett RM, Davis J. Lactoferrin interacts with deoxyribonucleic acid: a preferential reactivity with double-stranded DNA and dissociation of DNA-anti-DNA complex. J Lab Clin Med 1982; 99: 127-38.
13. Yamauchi K, Wakabayashi H, Shin K, Takase M. Bovine lactoferrin: benefits and mechanism of action against infections. Biochem Cell Biol 2006; 84: 291-6.
14. Connely OM. Antiinflammatory activities of lactoferrin. J Am Coll Nutr 2001; 20: 389-95.
15. Kalmar JR, Arnold RR. Killing of Actinobacillus actinomycetemcomitans by human lactoferrin. Infect Immun 1988; 56: 2552-2557.
16. Baker EN. Structure and reactivity of transfer­rins. Advances in Inorganic Chemistry 1994; 41: 389-463.
17. Zarember KA, Sugui JA, Chang YC, Kwon-Chung KJ, Gallin JI. Human polymorpho nuclear leukocytes inhibit Aspergillus fumigates conidial growth by lactoferrin-mediated iron depletion. J Immunol 2007; 178: 6367-73.
18. Baker EN, Baker HM. Lactoferrin molecular structure, binding properties and dynamics of lactoferrin. Cell Mol Life Sci 2005; 62: 2531-9.
19. Utsugi T, Schroit AJ, Connor J, Bucana CD, Fidler IJ. Elevated expression of phosphatidyl serine in the outer membrane leaflet of human tumor cells and recognition by activated human blood monocytes. Cancer Res 1991; 51: 3062-3066.
20. Bellamy W, Takase M, Wakabayashi H. Antibacterial spectrum of lactoferricin B, a potent bactericidal peptide derived from the N-terminal region of bovine lactoferrin. J Appl Bacteriol 1992; 73:472-479.
21. Kanyshkova TG, Babina SE, Semenov DV, Isaeva N, Valssov AV, Neustroev KN. Multiple enzymatic activities of human milk lactoferrin. Eur J Biochem 2003; 270: 3353-61.
22. Bailey TL, Boden M, Buske FA, Frith M, Grant CE, Clementi L, Ren J, Li WW, Noble WS. MEME SUITE: tools for motif discovery and searching. Nucleic Acids Res 2009; 37: 202-208.
23. Rost B. protein secondary structure prediction continues to rise. J Struct Biol 2001; 134: 204-218
24. Yamauchi K, Tomita M, Giehl TJ, Ellison RT. Antibacterial activity of lactoferrin and a pepsin derived lactoferrin peptide fragment. Infect Immun 1993; 61: 719-728.
25. Felsenstein J. Confidence limits on phylogenies: An approach using the bootstrap. Evolution 1985; 39: 783–791
26. Wang L, Brown SJ. BindN: a web-based tool for efficient prediction of DNA and RNA binding sites in amino acid sequences. Nucleic Acids Res 2006; 34: 243-248.
27. Lambert LA, Perri H, Meehan TJ.  Evolution of duplications in the transferrin family of proteins. Comp Biochem Physiol B Biochem Mol Biol 2005; 140: 11-25.
28. Marra AK, Jenssenb H, RoshanMoniria M, Hancock b REW, Pante N. Bovine lactoferrin and lactoferricin interfere with intracellular trafficking of Herpes simplex virus-1. Biochimie 2009; 91: 160-164.
29. Yi M, Kaneko S, Yu DY, Murakami S. Hepatitis C virus envelope proteins bind lactoferrin. J Virol 1997; 71: 5997-6002.
30. Geerts MEJ, Van Heen HA, Mericskay M, De Boer HA, Nuijens JH. N-terminal stretch Arg2, Arg3, Arg4 and Arg5 of human lactoferrin is essential for binding to heparin, bacterial lipopolysaccharide, human lysozyme and DNA. Biochem J 1997; 328: 145-151.
31. He J., Furmanski p. Sequence specificity and tran­scriptional activation in the binding of lactoferrin to DNA. Nature 1995; 373: 721–724.
32. Parillo JE. Pathogenetic mechanisms of septic shock. N Engl J Med 1993; 328:1471–1477.
33. Shin K, Wakabayashi H, Yamauchi K, Yaeshima T, Iwatsuki K. Recombinant Human Intelectin Binds Bovine Lactoferrin and Its Peptides. Biol Pharm Bull 2008; 3:1605-1608.
34. Ohashi A, Murata E. New functions of lactoferrin and β-casein in mammalian milk as cysteine protease inhibitors. Biochemical and Biophysical Research Communications 2003; 306: 98–103.
35. Orla M, Conneely PhD. Anti-inflammatory Activities of Lactoferrin. J Amer Col Nutri. 2001; 5: 389–395.