Intraparticle Macromolecular Migration Alters the Structure and Function of Proteins Reversibly Immobilized on Porous Microbeads

• Published in: Advanced materials interfaces Vol. 9; no. 18
• Main Authors: Diamanti, Eleftheria, Arana‐Peña, Sara, Ramos‐Cabrer, Pedro, Comino, Natalia, Carballares, Diego, Fernandez‐Lafuente, Roberto, López‐Gallego, Fernando
• Format: Journal Article
• Language: English
• Published: Weinheim John Wiley & Sons, Inc 01-06-2022
• Subjects: Biomedical materials; bio‐interfaces; Enzymes; Fluid dynamics; Fluid flow; Fluorescence; Immobilization; lipases; Nanoparticles; protein immobilization; protein storage; Proteins; single‐particle analysis
• ISSN: 2196-7350; 2196-7350
• DOI: 10.1002/admi.202200263

While migration of reversibly immobilized proteins across the volume of supports is investigated in conditions where an external force is applied or under fluid flow conditions, their passive migration upon sample storage and its effect on the protein functionality remain unexplored. Understanding such intraparticle macromolecular migration is essential to develop protein functionalized biomaterials with a longer life span. This work investigates the spatiotemporal migration of His‐tagged immobilized fluorescent proteins inside porous agarose microbeads under different storage conditions. A tool that assesses the intraparticle protein migration across the surface of the porous supports is developed. Differences in migration patterns between different proteins suggest that binding dynamics between proteins and their supports play a key role in their migration. The effect of macromolecular migration on the functional and structural properties of bound proteins and enzymes is also explored. Therefore, single‐particle measurements to understand how the migration process affects the functionality of immobilized enzymes are performed. Evaluating protein migration and understanding the reason behind such phenomena allows gaining control over immobilization processes and design immobilization chemistries that either prevent or promote intraparticle macromolecular diffusion upon storage, depending on the desired final application.
Spatiotemporal resolution studies reveal passive migration of macromolecules in 3D porous surfaces. Protein intraparticle passive migration, at the interface with solid surfaces, is demonstrated and shines light on the binding dynamics that rule out that phenomenon. Migration alters the structural and functional properties of immobilized proteins, including a case study for an industrially relevant enzyme.