Small Angle Neutron and X-Ray Scattering of Plasma Glycoprotein Interactions with Lipid Membranes

Abstract

Fibrinogen and Alpha 1-Antitrypsin (A1AT) are plasma glycoproteins with different, but specific functions. A1AT has been shown to have protective roles of lung cells against emphysema, while fibrinogen is a major factor in the blood clotting process. Most known glycoproteins have been shown to play a role in cellular interactions but the exact role of the glycan chains is still under investigation. Previous electrophysiological measurements show that A1AT has a strong affinity to lipid bilayers, perturbing the function of ion channels present in the membrane. These observed protein-membrane and protein-protein interactions in solution were studied using contrast-matching small-angle neutron scattering (SANS), small angle x-ray scattering (SAXS) and dynamic light scattering (DLS). To establish a structural reference point for each protein in solution, a series of polyethelene glycol-1,500 MW (PEG-1500) induced osmotic stress measurements were performed. The following D2O:H2O contrast matches were used for SANS: 40% for proteins, 15% for lipids and 16% for PEG-1500. Radius of gyration (Rg) approximations using Guinier analysis of A1AT in solution show a structural phase transition when the concentration of PEG-1500 is between 33% and 36% by volume. Significant structural changes were also observed for fibrinogen when the concentration of PEG-1500 was above 40% by volume. These structural changes were compared with changes observed when A1AT was in the presence of three different lipid membranes: POPC, POPS and DLPC and when fibrinogen undergoes polymerization. Bragg peaks produced by lipid membranes show that A1AT interacts with unilamellar vesicles. DLS was used to find suitable concentrations of fibrinogen (4 mg/ml) and thrombin (0.01 units/ml) that would yield a significant signal-to-noise ratio for SANS experiments (or approximately 90 minutes of reaction time). SANS time-resolved measurements show that fibrin structure is affected by its polymerization rate.