Effective Immobilization of hnRNPA2B1 Protein in a PEI Layer on a QCM Gold Electrode

RNA-binding proteins (RBPs) play a crucial role in RNA metabolism, influencing processes like transcription, splicing, transport, and stability, as well as cell proliferation and immune responses. Their links to diseases, such as cancer and neurological disorders, make them prime candidates for therapeutic targeting. Among these, heterogeneous nuclear ribonucleoprotein A2B1 (hnRNPA2B1) is notable for its regulation of gene expression and involvement in telomere maintenance and DNA repair. Its activity in various cancers and neurodegenerative diseases positions it as a promising target for drug development. The quartz crystal microbalance (QCM) method offers an efficient alternative to traditional binding affinity assessments such as spectroscopy, allowing experiments with minimal reagents and without extensive modifications. A key to effective QCM analysis is immobilization of the target protein to prevent denaturation. This study outlines a strategy for immobilizing hnRNPA2B1 onto a gold electrode using a polyethylenimine (PEI) layer. Adsorption processes and stability were monitored via frequency shift (Δf/n) and dissipation change (ΔD/n) measurements. The results showed that hnRNPA2B1's adsorption on branched PEI resulted in weak binding interactions, while adsorption on a linear PEI layer led to a negative frequency shift of -21 Hz. Increasing the ionic strength to 0.1 mM significantly enhanced protein adsorption (Δf/n = -69 Hz). These findings emphasize the role of the PEI layer structure in optimizing protein immobilization, paving the way for further exploration of RBPs and their ligands.