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Biofilm-related infections are associated with high mortality and morbidity combined with increased treatment costs. Traditional antibiotics are becoming less effective due to the emergence of drug-resistant bacterial strains. The need to treat biofilms on medical implants is particularly acute, and one persistent challenge is selectively directing nanoparticles to the biofilm site. Here, we present a protein-based functionalization strategy that targets the extracellular matrix of biofilms. The engineered protein combines the Staphylococcus epidermidis autolysin R2ab domain with a gold-binding GB3 domain, directing nanoparticles specifically to bacterial cell wall components (lipoteichoic acid and wall teichoic acid) that are absent in mammalian tissues. This fusion protein is applied to a gold nanoparticle (AuNP) core, along with elastin-like polypeptides (ELPs), which generate a robust photothermal response. The engineered particles exhibit exceptional biocompatibility, including low protein corona formation, minimal macrophage uptake, and hemocompatibility, while maintaining selective biofilm targeting. The photothermal conversion can be modulated by changing the ELP transition temperature, and the functionalized AuNPs strongly interact with biofilms under static and flow conditions without significantly binding to serum-coated surfaces. Near-infrared laser irradiation resulted in a 10,000-fold improvement in killing efficiency compared to untreated controls (p < 0.0001). The targeting strategy utilized here represents a versatile approach to targeting drug-resistant infections and could be readily expanded to other bacterial pathogens and anti-biofilm nanoparticle platforms. 

When nanoparticles and nanoplastics enter biological fluids, their surfaces are rapidly coated with proteins, forming a corona that governs biological responses. However, understanding protein- surface interaction energetics remains a significant challenge. Here, we examine how protein charge distribution affects adsorption to polystyrene nanoparticles (PSNPs) by generating a series of lysine-to-alanine variants of the GB3 protein. Using isothermal titration calorimetry (ITC), we found that the K19A variant binds most strongly to both non-functionalized and carboxylate- functionalized PSNPs. ITC thermograms indicate that K19A forms a stable monolayer, while other variants exhibit multilayer adsorption. We hypothesize that removing lysine at position 19 creates a flatter, more neutral interaction surface that promotes efficient initial binding. Fluorescence denaturation experiments show that PSNPs destabilize GB3 protein variants, and binding correlates strongly with protein unfolding (r = 0.82, p < 0.01 for COOH-PSNPs and r = 0.76, p < 0.03 for non-functionalized PSNPs). These results reveal how protein stability and charge distribution shape adsorption thermodynamics, offering a framework for predicting protein-surface interactions.