Solution structure and backbone dynamics of human epidermal-type fatty acid-binding protein (E-FABP)

Abstract

Human epidermal-type fatty acid-binding protein (E-FABP) belongs to a family of intracellular 14-15 kDa lipid-binding proteins, whose functions have been associated with fatty acid signalling, cell growth, regulation and differentiation. As a contribution to understanding the structure-function relationship, we report in the present study features of its solution structure and backbone dynamics determined by NMR spectroscopy. Applying multi-dimensional high-resolution NMR techniques on unlabelled and 15N-enriched recombinant human E-FABP, the 1H and 15N resonance assignments were completed. On the basis of 2008 distance restraints, the three-dimensional solution structure of human E-FABP was subsequently obtained (backbone atom root-mean-square deviation of 0.92+/-0.11 A; where 1 A=0.1 nm), consisting mainly of 10 anti-parallel beta-strands that form a beta-barrel structure. 15N relaxation experiments (T1, T2 and heteronuclear nuclear Overhauser effects) at 500, 600 and 800 MHz provided information on the internal dynamics of the protein backbone. Nearly all non-terminal backbone amide groups showed order parameters S(2)>0.8, with an average value of 0.88+/-0.04, suggesting a uniformly low backbone mobility in the nanosecond-to-picosecond time range. Moreover, hydrogen/deuterium exchange experiments indicated a direct correlation between the stability of the hydrogen-bonding network in the beta-sheet structure and the conformational exchange in the millisecond-to-microsecond time range. The features of E-FABP backbone dynamics elaborated in the present study differ markedly from those of the phylogenetically closely related heart-type FABP and the more distantly related ileal lipid-binding protein, implying a strong interdependence with the overall protein stability and possibly also with the ligand-binding affinity for members of the lipid-binding protein family.

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

“…Previous studies with various i-LBPs have revealed a correlation between ligand-binding affi nity and overall structural stability for several members of this protein family ( 21,36 ). In order to determine whether such a correlation also applies to the cellular retinol carriers, we have now nearly identical to those at pH 6.0 (differences р 0.02 ppm), as shown in Table 2 .…”

“…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 ).…”

“…NMR investigations of various i-LBPs have revealed that their backbone dynamics are correlated with the respective binding properties ( 20,21 ). 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 ).…”

Nuclear Magnetic Resonance Structure-Based Epitope Mapping and Modulation of Dust Mite Group 13 Allergen as a Hypoallergen

“…Three-dimensional structures of these different FABPs have been extensively studied and found to be highly conserved, although their sequence similarities could vary between 20 and 70%. These proteins adopt a mixed ␣-␤ structure, with a typical ␤ barrel made up of 10 antiparallel ␤ strands and covered with a pair of ␣ helices at one end (15)(16)(17).…”

Proteomics and Esophageal Cancer

“…It was proposed that they play a role in cellular lipid uptake and transport, metabolic pathway, and regulation of protein metabolism. Downregulation of E-FABP in has been reported for ESCC [46].…”