Numerous applications in analytical chemistry, bio-catalysis, biotechnology and in various medical devices are faced with an inherent problem relating to protein immobilization at interfaces. Basically, for successful application, the immobilization should not alter the protein conformation and maintain its bioactivity. The perfect method should also be unspecific with respect to the surface chemistry, surface geometry, and the protein to be immobilized. To this end, it is promising to adapt the so-called layer-by-layer (LbL) assembly of polyelectrolyte (PE), to the protein immobilization. This method consists in the alternate adsorption of oppositely charged PE, which results in a PE multilayer. By alternating the adsorption of a protein with a PE, it is, in theory, possible to construct a multilayer that immobilizes the protein. Nevertheless, integrating proteins in LbL thin films is very challenging due to their low conformational entropy, heterogeneous spatial distribution of charges, and polyampholyte nature.
Protein−polyelectrolyte complexes (PPCs) are promising building blocks for LbL construction owing to their standardized charge and polyelectrolyte corona. Unfortunately, the true advantage that they bring compared to bare protein integration have not been shown yet.
To this note, a team of researchers at UCLouvain led by Professor Christine Dupont-Gillain from the Institute of Condensed Matter and Nanosciences in Belgium demonstrated that the electrical charge of a protein could be standardized by a polyelectrolyte (PE) and that the resulting PPCs could be used to obtain sustainable multilayer growth. In particular, they showed that the multilayer construction using PPCs could be based on PE−PE interactions only and as such would allow the construction method to be generalized to all proteins. Their work is currently published in the research journal, ACS Nano.
In brief, their research team prepared PPCs through the complexation of lysozyme with low molar mass poly(styrene sulfonate) (PSS) in different conditions of pH and ionic strength. Next, the PPCs size and electrical properties were investigated, and the forces driving complexation were elucidated, in the light of computations of PE conformation, with a view to further unravel LbL construction mechanisms. Lastly, quartz crystal microbalance and atomic force microscopy were used to monitor the integration of PPCs compared to the one of bare protein molecules in LbL assemblies, and colorimetric assays were performed to determine the protein amount in the thin films.
The authors observed that the layers built with PPCs showed higher protein contents and hydration levels. In addition, the researchers noted that LbL construction with PPCs mainly relied on standard PE−PE interactions, i.e. independent of the charge state of the protein, in contrast to classical bare protein assembly with polyelectrolytes.
In summary, in the frame of the PhD thesis of Aurélien vander Straeten, the team of researchers presented the standardization of the electrical charge of a protein using polyelectrolyte, where the resulting PPCs could be used to obtain sustainable LbL growth. In general, it was seen that the balance between intramolecular electrostatic repulsion within PSS and electrostatic attraction between PSS and lysozyme was responsible for PPCs formation. Altogether, their observations considerably simplify the incorporation of proteins in multilayers, which will be beneficial for biosensing, heterogeneous bio-catalysis, biotechnologies, and medical applications that require active proteins to be immobilized on various surfaces.
Aurelien vander Straeten, Anna Bratek-Skicki, ́ Alain M. Jonas, Charles-André Fustin, Christine Dupont-Gillain. Integrating Proteins in Layer-by-Layer Assemblies Independently of their Electrical Charge. ACS Nano 2018, volume 12, page 8372−8381.Go To ACS Nano