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Biointerphases

Journal for Biophysical Chemistry

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Interfacial adsorption and denaturization of human milk and recombinant rice lactoferrin

Biointerphases volume 3, pages FB36–FB43 (2008)Cite this article

Abstract

Lactoferrin (LF) produced from recombinant technologies can achieve almost identical amino acid sequences and three-dimensional structures to those extracted from mammals, but differences often arise in the carbohydrate chains attached through N-glycosylation, with altered sizes, structures, and chemical nature. We compare the differences in solvation and interfacial adsorption from two samples, human milk lactoferrin (hLF) and recombinant rice lactoferrin (rLF). Lactoferrin is a bilobal protein with a molecular weight of about 80 kD. It has three N-glycosylation sites. Each of the three attached glycan chains on rLF contains seven to eight sugar groups. In comparison, each of the three glycan chains attached to hLF contains 12–13 sugar groups and is twice as long. The rLF meting point in 1 mg/ml aqueous solution (pH 7 phosphate buffer, I=20 mM was 43 °C from dynamic light scattering, compared to 53 °C for hLF, exhibiting the enhanced solvation and stability of hLF due to its longer carbohydrate side chains. Silicon oxide surfaces provided a model substrate for assessment of lactoferrin adsorption and comparison with other proteins. The time dependent interfacial adsorption studied by spectroscopic ellipsometry (SE) was characterized by a fast initial step followed by a slow relaxation process. In addition, the SE results revealed the persistently higher adsorption of rLF, again showing the effect of glycan side chains. In spite of the different adsorbed amounts, neutron reflection revealed similar interfacial structures of the adsorbed protein layers. At the low lactoferrin concentration around 10 mg/l, a flat-on molecular monolayer formed with both LF lobes attached to the SiO2 surface through electrostatic attraction. As the protein concentration increased, a secondary molecular layer further adsorbed to the first one and the attachment was again driven by electrostatic attraction. The intermixing between the globular lobes resulted in the dense packing in the middle 60 Å with some of the lobes projected toward the aqueous bulk solution.

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Authors and Affiliations

  1. Biological Physics Group, Schuster Building, The University of Manchester, Oxford Road, M13 9PL, Manchester, UK

    Fang Pan, XiuBo Zhao, Thomas A. Waigh & Jian R. Lua

  2. S.I.F.I. S.p.A., Via Ercole Patti, 36-95020 Aci S. Antonio-Lavinaio, Catania, Italy

    Fausto Miano

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  1. Fang Pan
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  2. XiuBo Zhao
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  3. Thomas A. Waigh
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Pan, F., Zhao, X., Waigh, T.A. et al. Interfacial adsorption and denaturization of human milk and recombinant rice lactoferrin. Biointerphases 3, FB36–FB43 (2008). https://doi.org/10.1116/1.2965135

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  • DOI: https://doi.org/10.1116/1.2965135