Fatty acid binding to ovalbumin studied by EPR

Abstract

Serum albumins (SAs) act as main carriers of various endogenous and exogenous compounds, including non-esterified fatty acids (FAs), in the blood plasma of mammals. Avian SAs exhibit certain similarities and moderate sequence identity to mammalian SAs, i.e. human and bovine (HSA and BSA). Chicken SA (α-livetin) was found to transport FAs like the mammalian, although with lower affinity towards long-chain FAs. This protein can be found in the hens blood serum and the egg yolk, but not in the egg white. Ovalbumin (OVA), the albumin from hen egg white, accounts for more than half of the total albumen protein content, while in the egg yolk appears only in its dephosphorylated form, presumably serving as the nutrition source for the growing embryo. OVA belongs to the serpin superfamily, despite lacking any protease inhibitory activity. Although it is the first protein isolated in pure form from egg white, its physiological function has not yet fully been resolved. It is mainly proposed that OVA serves as a transport and storage protein for metal ions and amino acids. The aim of this study was to determine whether OVA binds long-chain FAs, and to compare it with HSA and BSA, using electron paramagnetic resonance spectroscopy (EPR) spin labeling, a technique well established in the characterization of SA binding properties. The proteins were labeled with 5-, and 16-doxyl-stearic acids (5-DS and 16-DS). The EPR spectral analysis revealed strong binding of 5-DS and 16-DS to both HSA and BSA, and only slight immobilisation in a 1:1 molar ratio solution with OVA. Most probably, the observed restricted motion of FAs is the consequence of their surface interactions with OVA residues, rather than specific binding to the protein, as observed for SAs. These results indicate that OVA lacks a FA binding site, unlike SAs which contain multiple high-affinity FA binding sites. Moreover, it may be concluded that FA binding and transport is not a primary physiological function of OVA.