DNA, oligosaccharides, and mononucleotides stimulate oligomerization of human lactoferrin

Abstract

Using small-angle X-ray scattering (SAXS), light scattering (LS), and soft laser ablation we have shown that lactoferrin (LF) in solution at neutral pH is oligomerized in the absence of salt or at physiological salt concentrations. The level of oligomerization depends on the concentration of LF, KCl or NaCl, and on the duration of the protein storage in solution. At the concentrations comparable with those in human milk (1-6 mg/ml), the average radius of gyration (R(g)) values of LF can attain 400-480 A, while fresh solution of previously lyophylized LF demonstrate a lower average R(g) (50-100 A), and R(g) value characterizing the LF monomer formed at 1 M NaCl is 26.7 A. The addition of oligonucleotides, oligosaccharides, or mononucleotides to LF in the presence or in the absence of KCl with different level of initial oligomerization accelerates the oligomerization rate and increases the R(g) values up to approximately 600-700 A, which correspond to associates containing ten or more protein molecules. During gel filtration on Sepharose 4B, high-degree LF oligomers dissociate nearly completely forming different degraded complexes, but in some cases it is possible to reveal small amount of a decamer. A possible role for oligomerization of LF, a highly polyfunctional protein, for its different biological activities is discussed.

Identification of Unprecedented Anticancer Properties of High Molecular Weight Biomacromolecular Complex Containing Bovine Lactoferrin (HMW-bLf)

“…In order to identify the homogeneity of HMW-bLf, analysis of the components of HMW-bLf was further carried out by dissociating the complex into its dimeric and monomeric forms in the presence of 1 M NaCl as described earlier for hLf [11]. The study reported that hLf oligomers dissociate fast and almost completely to monomers in the presence of high concentrations (≥1.0 M) of Na + or K + .…”

“…Briefly, 3 mg mL −1 of HMW-bLf was dissolved in 50 mM Tris HCl (pH 7.5) containing 1 M NaCl and incubated for 1 day at 37°C. SDS-PAGE was carried out to analyze the dissociation of HMW-bLf complex into its components [11]. Western blotting was carried out to analyze the obtained protein bands by goat anti- bLf (Bethyl Laboratories) diluted in the ratio of 1∶1000 to confirm the presence of lactoferrin.…”

“…Interestingly, the addition of oligonucleotides, oligosaccharides, or mononucleotides to hLf in the presence or absence of KCl accelerated the oligomerization rate leading to the formation of associates containing ten or more protein molecules. The presence of ions, ATP, NAD, nucleotides, DNA or polysaccharides can further effect the self-association of Lf molecule under physiological conditions [11]. These findings suggest the importance of oligomerization for the multifunctional activities of Lf during host–pathogen interactions, and also in targeting cellular and molecular components of disease signaling pathways.…”

“…Recently, by employing different techniques such as gel filtration, soft laser ablation, small-angle X-ray scattering (SAXS), and light scattering (LS), hLf has been reported to oligomerize into several high molecular weight (HMW) aggregates (70 kDa–800 kDa). The level of oligomerization was reported to depend on the concentrations of Lf, KCl, NaCl and also on the duration of the protein storage in solution [11]. Interestingly, the addition of oligonucleotides, oligosaccharides, or mononucleotides to hLf in the presence or absence of KCl accelerated the oligomerization rate leading to the formation of associates containing ten or more protein molecules.…”

Cloning, expression and characterization of Kunming mice lactoferrin and its N-lobe

“…Some studies concluded that iron binding activity and restricting iron metabolism are the foundation of inhibiting the growth of microbe (Caraher et al 2007;Zarember et al 2007). Besides, some reports demonstrated that the ability to bind to other macromolecules (proteins, DNA) (Baker and Baker 2009;Nevinskii et al 2009) may also play important role in the many functions. The modeled distribution of surface charge (Fig.…”