Characterization of the Vitamin E-Binding Properties of Human Plasma Afamin

Abstract

Human plasma afamin, the fourth member of the albumin gene family, is shown to be a specific binding protein for vitamin E. A radio ligand-binding assay followed by Scatchard and Hill analysis are used to show that afamin has a binding affinity for both alpha-tocopherol and gamma-tocopherol, two of the most important forms of vitamin E, in vitro. The binding-dissociation constant was determined to be 18 microM, indicating that afamin plays a role as vitamin E carrier in body fluids such as human plasma and follicular fluid under physiological conditions. Additionally, we demonstrate that afamin has multiple binding sites for both alpha- and gamma-tocopherol. Due to the large binding capacity of afamin for vitamin E, it might take over the role of vitamin E transport in body fluids under conditions where the lipoprotein system is not sufficient for vitamin E transport. To confirm the experimental results, we performed homology modeling and docking calculations on the predicted tertiary structure, which showed coincidence between calculated and in vitro results.

Afamin and Apolipoprotein A-IV: Novel Protein Markers for Ovarian Cancer

“…It will also be interesting to look at possible accumulations of both proteins in the ascites fluid of the tumor. In this context, it is noteworthy to speculate about possible antioxidative properties of both proteins (18,24) and their subsequent putative response to tumor growth.…”

“…Afamin was quantified with a double-antibody sandwich ELISA using an affinity-purified polyclonal antibody for coating 96-well microtiter plates and the peroxidase-conjugated monoclonal antibody N13 for detection (16,18). The coating antibody had been previously biotinylated and bound to streptavidincoated plates (MicroCoat).…”

“…Statistically significant decreased serum concentrations of afamin were found in 57 ovarian cancer patients compared with 60 patients with benign gynecologic conditions, 26 patients with borderline ovarian cancer, and 39 healthy controls. Afamin is a previously described member of the albumin multigene family with vitamin E-binding properties shown in vitro (16)(17)(18). In contrast to other family members, afamin is highly glycosylated and exerts a molecular weight of 87 kDa.…”

Complexity of vitamin E metabolism

“…However, in body fluids with low lipoprotein concentrations, such as follicular fluids, an alternative carrier protein for vitamin E, namely afamin, has been described [87][88][89] . Afamin is a liver-derived plasma glycoprotein and a member of the albumin protein family [90][91][92] ; therefore it is also called α-albumin [93] .…”

“…Afamin is partly (13%) bound to plasma lipoproteins [88] , but circulates mostly in free form. Since afamin has 18 predicted binding sites for vitamin E and shows binding affinity for both α-TOH and γ-TOH, it has been suggested to be an alternative vitamin E transporter in body fluids under conditions where the lipoprotein system is not sufficient for vitamin E transport [89] . Originally, vitamin E was discovered as a resorption-gestations factor in female rats [2,94] .…”