Spectroscopic and Molecular Modeling Investigation on the Interaction between Folic Acid and Bovine Lactoferrin from Encapsulation Perspectives

Abstract

The impact of thermal treatment on the ability of lactoferrin (FL) to bind folic acid (FA) was investigated by employing fluorescence spectroscopy, molecular dynamics and docking tests. The structural and conformational particularities of LF upon heating at 80 °C and 100 °C were first estimated based on the intrinsic fluorescence changes in respect to the native protein. The emission spectra indicated gradual unfolding events accompanied by Trp exposure with increasing temperature. In agreement with the experimental results, molecular modeling investigations showed that the secondary and tertiary structure of LF are slightly affected by the thermal treatment. Some minor unfolding events related particularly to the α-helical regions of LF were observed when the temperature increased to 100 °C. The LF fluorescence quenching upon FA addition indicated that a static mechanism stands behind LF-FA complex formation. Regardless of the simulated temperature, the hydrogen bonds played an important role in regulating the interaction between the protein and ligand. FA binding to LF equilibrated at different temperatures occurred spontaneously, and all complexes displayed good thermodynamic stability. The obtained results support the suitability of LF as biocompatible material, for obtaining micro- and nanoparticles for delivery of dietary supplements or for enhancing the functionality of target delivery systems.

Keywords: dietary; folic acid; functional; interactions; lactoferrin; molecular modeling.

Figure 1

The fluorescence spectra of LF in the absence of the quencher.

Figure 2

The fluorescence spectra of the interaction between LF thermally treated at 25 °C (a); 80 °C (b) and 100 °C (c) and FA. The FA concentration (from A–F) varied from 0 to 11.32 × 10⁻⁸ M.

Figure 3

Stern-Volmer plots for the interaction of folic acid with heat-treated lactoferrin.

Figure 4

Superposition of the lactoferrin-folic acid complexes involving the protein pre-heated at different temperatures. The protein is represented in New Cartoon style. Domains N-1 (amino acids 1 to 90 and 251 to 333) and N-2 (amino acids 91 to 250) of the N lobe are represented in blue and lime, respectively. Domains C-1 (amino acids 345 to 431 and 593 to 676) and C-2 (amino acids 432 to 592) of the C lobe are represented in orange and cyan, respectively. The folic acid of the complexes involving proteins equilibrated at 25 °C, 80 °C and 100 °C, is represented in van de Waals style in mauve, violet and red, respectively.