New insights into intracellular lipid binding proteins: The role of buried water

Abstract

The crystal structures of most intracellular lipid binding proteins (LBPs) show between 5 and 20 internally bound water molecules, depending on the presence or the absence of ligand inside the protein cavity. The structural and functional significance of these waters has been discussed for several LBPs based on studies that used various biophysical techniques. The present work focuses on two very different LBPs, heart-type fatty acid binding protein (H-FABP) and ileal lipid binding protein (ILBP). Using high-resolution nuclear magnetic resonance spectroscopy, certain resonances belonging to side-chain protons that are located inside the water-filled lipid binding cavity were observed. In the case of H-FABP, the pH-and temperaturedependent behavior of selected side-chain resonances (Ser82 OgH and the imidazole ring protons of His93) indicated an unusually slow exchange with the solvent, implying that the intricate hydrogen-bonding network of amino-acid side-chains and water molecules in the protein interior is very rigid. In addition, holo H-FABP appeared to display a reversible self-aggregation at physiological pH. For ILBP, on the other hand, a more solvent-accessible protein cavity was deduced based on the pH titration behavior of its histidine residues. Comparison with data from other LBPs implies that the evolutionary specialization of LBPs for certain ligand types was not only because of mutations of residues directly involved in ligand binding but also to a refinement of the internal water scaffold.

New insights on the protein-ligand interaction differences between the two primary cellular retinol carriers

“…A critical factor for the structural stability of i-LBPs is a cluster of structural water molecules inside the binding cavity, which developed during the course of an evolutionary specialization of this protein family ( 23 ). These water molecules form an intricate network of hydrogen bonds among each other and with several side chains in the protein interior, thereby strengthening the overall protein structure as previously demonstrated by H/D exchange experiments performed with other i-LBPs ( 21 ).…”

“…The substitutions L29T and P38Q, involving residues in contact with retinol, were discussed in the previous section; the substitution A121C is located in the ␤ -turn IJ, where the neighboring residue 122 can form a salt bridge only in CRBP-I. The four remaining nonconservative substitutions F4Q, I51T, L74T, and C82V, all involving the internal network of structural water molecules described for heart fatty acid binding protein (FABP) ( 23 ), could be of par- …”

“…The nonconservative substitutions I51T and L74T were already mentioned with respect to the internal network of structural water molecules, which acts in several members of the i-LBP family as a kind of hydration shell for the hydrophobic residues inside the ligand-binding cavity ( 23 ). The presence of Met (in CRBP-I) instead of either Leu (in rat CRBP-II) or Val (in human CRBP-II) at position 62 is of the homologs: two in CRBP-I between Glu-41 and Arg-52 as well as between Lys-101 and Glu-122, and one in CRBP-II between Lys-50 and Asp-63 ( Fig.…”

“…It appears that the i-LBP family evolved from a multifaceted ancestral precursor into highly specialized lipid carriers that transport either retinoids or fatty acids through the cytosol of various tissues ( 22,23 ).…”

Structural and dynamic roles of permanent water molecules in ligand molecular recognition by chicken liver bile acid binding protein

“…These biopolymers contain several water molecules inside their binding site; the number is different in each specific macromolecule and even between the apo and the holo forms of the same protein Zanotti et al, 1992;Rich and Evans, 1996;Mesgarzadeh et al, 1998;Likic and Prendergast, 1999;Wiesner et al, 1999;Woolf and Tychko, 1999;Woolf et al, 2000;Lucke et al, 2002;van Gunsteren, 2002a, 2002b;Bottoms et al, (www.interscience.wiley.com) DOI:10.1002/jmr.908 Research Article 2006). Some of these water molecules are essential for the stabilization of the entire protein fold and/or for ligand binding and interaction selectivity (Lucke et al, 2002). For these reasons, when studying these proteins through computational methods, the binding site water molecules have to be treated in an explicit manner.…”