No ordinary proteins: Adsorption and molecular orientation of monoclonal antibodies

Abstract

The interaction of monoclonal antibodies (mAbs) with air/water interfaces plays a crucial role in their overall stability in solution. We aim to understand this behavior using pendant bubble measurements to track the dynamic tension reduction and x-ray reflectivity to obtain the electron density profiles (EDPs) at the surface. Native immunoglobulin G mAb is a rigid molecule with a flat, “Y” shape, and simulated EDPs are obtained by rotating a homology construct at the surface. Comparing simulations with experimental EDPs, we obtain surface orientation probability maps showing mAbs transition from flat-on Y-shape configurations to side-on or end-on configurations with increasing concentration. The modeling also shows the presence of β sheets at the surface. Overall, the experiments and the homology modeling elucidate the orientational phase space during different stages of adsorption of mAbs at the air/water interface. These finding will help define new strategies for the manufacture and storage of antibody-based therapeutics.

Differential Surface Adsorption Phenomena for Conventional and Novel Surfactants Correlates with Changes in Interfacial mAb Stabilization

“…For all the mAb concentrations studied, the surface tension decreases and reaches a quasi-equilibrium value as molecules adsorb to the surface. At long times, the tension continues to decrease and does not achieve an equilibrium value, indicating irreversible adsorption of mAbs upon adsorption due to unfolding or slow rearrangements of the adsorbed mAbs. , …”

“…This value is lower than the area per molecule for the maximum packing of the flat-on orientation (18,750 Å 2 ) but higher than that for the side-on (6875 Å 2 ) or end-on (8250 Å 2 ) orientation. Prior studies using homology modeling by our group have indicated that the adsorbed layer surface population could consist of a mixture of all three orientations (flat-on, side-on, and end-on) …”

“…Prior studies using homology modeling by our group have indicated that the adsorbed layer surface population could consist of a mixture of all three orientations (flat-on, side-on, and end-on). 52 Mixed Adsorbed Layers. The qualitative comparison of XRR profiles of protein concentration of 0.5 and 2.5 mg/mL with PS80 or FM1000 is discussed in detail (see Figures S4 and S5 for protein concentration results at 1.0 mg/mL with PS80 and FM1000, respectively, in the Supporting Information).…”

Versatile and Easily Designable Polyester-Laser Toner Interfaces for Site-Oriented Adsorption of Antibodies

“…Binding to a strongly hydrophobic surface entails the risk of protein unfolding and exposition of hydrophobic domains, resulting in a partial denaturation, which is a typical phenomenon observed for passive adsorption of antibodies [ 41 , 42 ]. The reported degree of Abs denaturation may reach about 50%, which derives from protein conformational changes and random immobilization, hindering solvent accessibility of binding sites [ 43 ]. It is however worth underlining that in our case this effect is fully compensated for by the benefits resulting from the increased surface area.…”

Degradation of lactoferrin caused by droplet atomization process via two-fluid nozzle: The detrimental effect of air–water interfaces

“…Specifically, the tendency to adsorb onto the interface (e.g., greater foaming ability) increases the level of denaturation. 31 In this aspect, the equilibrium surface tension of lactoferrin (or the surface activity of lactoferrin) is similar to iGg, 43 rhGH, 59 mAbs, 60 and lysozyme 61 (with an equilibrium surface tension of 20 µM ~ 55-60 mN/m). These are proteins known to degrade with exposure to the air-water interface through the aeration of the solution.…”