Two modes of fatty acid binding to bovine β‐lactoglobulin—crystallographic and spectroscopic studies

Abstract

Lactoglobulin is a natural protein present in bovine milk and common component of human diet, known for binding with high affinity wide range of hydrophobic compounds, among them fatty acids 12-20 carbon atoms long. Shorter fatty acids were reported as not binding to β-lactoglobulin. We used X-ray crystallography and fluorescence spectroscopy to show that lactoglobulin binds also 8- and 10-carbon caprylic and capric acids, however with lower affinity. The determined apparent association constant for lactoglobulin complex with caprylic acid is 10.8 ± 1.7 × 10(3) M(-1), while for capric acid is 6.0 ± 0.5 × 10(3) M(-1). In crystal structures determined with resolution 1.9 Å the caprylic acid is bound in upper part of central calyx near polar residues located at CD loop, while the capric acid is buried deeper in the calyx bottom and does not interact with polar residues at CD loop. In both structures, water molecule hydrogen-bonded to carboxyl group of fatty acid is observed. Different location of ligands in the binding site indicates that competition between polar and hydrophobic interactions is an important factor determining position of the ligand in β-barrel.

Structural and thermodynamic studies of binding saturated fatty acids to bovine β-lactoglobulin

“…The structure revealed that lactoglobulin has similar fold to retinol binding protein and with its eight-stranded ␤-barrel can be classified as a member of lipocalin family [4]. Since that, many crystal structures of liganded and unliganded LGB have been determined, among them high-resolution crystal structures of trigonal and orthorhombic lactoglobulin forms have been reported [5,6]. The LGB ability to bind aliphatic alkanes was first noticed by Wishnia and Pinder [7].…”

“…They also determined dissociation constants for LGB complexes with other saturated and unsaturated fatty acids [10]. In our previous studies [6], we showed that LGB could also form complexes with shorter fatty acids such as capric and caprylic acids.…”

“…It was also demonstrated that lactoglobulin complex with linoleic acid can be potentially used in colon cancer-targeted therapy [39]. Other potential applications of lactoglobulin complexes prompted us to undertake systematic structural and thermodynamic studies of fatty acids binding to lactoglobulin [6].…”

Effects of Encapsulated Fish Oil by Polymerized Whey Protein on the Textural and Sensory Characteristics of Low-Fat Yogurt

“…Some studies have shown that fat droplets may bind to β-lactoglobulin (the fi rst major protein in WPC) at two different sites. The fi rst site is the central calyx located on the β-sheet strands where most of the hydrophobic groups are located and the other is on the protein surface itself [Loch et al, 2011]. Heat t reatment and homogenization may help α-lactalbumin (the second major protein in WPC) obtain a more open structure due to electrostatic repulsive charges, which may help proteins to spread out across a fat droplet surface and enhances its droplet coverage [Zhai et al, 2012;L am & Ni ckerson, 2015a].…”

Computational study of ligand binding in lipid transfer proteins: Structures, interfaces, and free energies of protein‐lipid complexes

“…(i) Caprate (CAP) from its complex with maize nsLTP, PDB entry 1FK0, [11] (ii) laurate (LAU) from peach nsLTP structure, PDB entry 2ALG, [14] (iii) palmitate (PLM) from its complex with maize nsLTP, PDB entry 1MZM, [10] (iv) 9,12-linoleate (EIC) from its complex with bovine b-lactoglobulin, PDB entry 3QZK, [21] (v) a-linolenate (LNL) from its complex with maize nsLTP, PDB entry 1FK6, [11] (vi) prostaglandin B 2 (PG2) from its complex with wheat nsLTP, PDB entry 1CZ2, [16] and (vii) lysomyristoylphosphatidyl-choline (LPC) from its complex with wheat nsLTP, PDB entry 1BWO. [12] Except (vi), which is a solution NMR structure, the geometries correspond to X-ray crystal structures.…”