Ligand entry into the calyx of β‐lactoglobulin

Abstract

Although the thermodynamic principles that control the binding of drug molecules to their protein targets are well understood, the detailed process of how a ligand reaches a protein binding site has been an intriguing question over decades. The short time interval between the encounter between a ligand and its receptor to the formation of the stable complex has prevented experimental observations. Bovine β-lactoglobulin (βlg) is a lipocalin member that carries fatty acids (FAs) and other lipids in the cellular environment. Βlg accommodates a FA molecule in its highly hydrophobic cavity and exhibits the capability of recognizing a wide variety of hydrophobic ligands. To elucidate the ligand entry process on βlg, we report molecular dynamics simulations of the encounter between palmitate (PA) or laurate (LA) and βlg. Our results show that residues localized in loops at the cavity entrance play an important role in the ligand penetration process. Analysis of the short-term interaction energies show that the forces operating on the systems lead to average conformations very close to the crystallographic holo-forms. Whereas the binding free energy analysis using the molecular mechanics Generalized Born surface area method shows that these conformations were thermodynamically favorable.

Fatty acid binding into the highest affinity site of human serum albumin observed in molecular dynamics simulation

“…For binding events of long duration (t bind > 1 ns), the fatty acid molecule spends a relatively long time probing the entry of the protein pocket. This behavior is in line with observations of other fatty acid binding events in MD simulations [15,16,18].…”

“…Differences with penetration times of $1 ns previously reported for other protein-fatty acid binding processes [15,18] can be due, at least in part, to differences in the conformation of the binding sites (e.g., geometry, hydration properties, composition in terms of amino acid residues, etc.). Nevertheless, and more significantly, our results indicate that the variability due to the intrinsic stochasticity of the process likely plays a major role.…”

“…Unfortunately, microscopic details concerning the dynamics of the lipid binding are currently not easily accessible to the experiment. Molecular dynamics (MD) 1 represents an interesting tool to tackle this issue [13,14] and has already been successfully employed in a few cases [15][16][17][18][19][20].…”

“…Bello and García-Hernández performed a total of 20 unbiased MD runs to explore the binding of lauric and palmitic acid to b-lactoglobulin [18], starting with the ligand outside the protein binding site. The ligand was observed to have at least a partial penetration in approximately half of the simulations, and ended up forming a stable complex in three instances.…”

Factors affecting the interactions between beta-lactoglobulin and fatty acids as revealed in molecular dynamics simulations

“…1 Bovine BLG is a relatively small protein of 162 amino acid residues, with a molecular weight of 18.4 kDa. 2 There are several genetic variants of BLG, with the A and B variants being the most common which differ by two amino acids at positions 64 and 118. 3 These two variants both have nine beta-strands which form a flattened and conical barrel, called a calyx (Fig.…”