Activity Study of Self-Assembled Proteins on Nanoscale Diblock Copolymer Templates

Abstract

Novel methods for affixing functional proteins on surfaces with high areal density have the potential to promote basic biological research as well as various bioarray applications. The use of polymeric templates under carefully balanced thermodynamic conditions enables spontaneous, self-assembled protein immobilization on surfaces with spatial control on the nanometer scale. To assess the full potential of such nanometer-scale protein platforms in biosensing applications, we report for the first time the biological activity of proteins on diblock copolymer platforms. We utilized horseradish peroxidase, mushroom tyrosinase, enhanced green fluorescent protein, bovine immunoglobulin G, fluorescein isothiocyanate conjugated anti-bovine IgG, and protein G as model systems in our protein activity studies. When specific catalytic functions of HRP and MT, immobilized on selective domains of microphase-separated PS-b-PMMA, are evaluated over a long period of time, these enzymes retain their catalytic activity and stability for well over 3 months. By performing confocal fluorescence measurements of self-fluorescing proteins and interacting protein/protein systems, we have also demonstrated that the binding behavior of these proteins is unaffected by surface immobilization onto PS-b-PMMA diblock copolymer microdomains. Our polymer platforms provide highly periodic, high-density, functional, stable surface-bound proteins with spatial control on the nanometer scale. Therefore, our diblock copolymer-guided protein assembly method can be extremely beneficial for high-throughput proteomic applications.

Controlling Protein Adsorption through Nanostructured Polymeric Surfaces

“…As presented in numerous studies, various proteins exhibit biased adsorption to one of the polymeric nanodomains (Figure 7a). [124][125][126][127] For instance, fibrinogen, horseradish peroxidase, or albumin, was found to selectively adsorb onto the PS nanodomains of the diblock copolymer surfaces (Figure 7b). [41] Furthermore, as shown in Figure 7c, protein adsorption is highly favored on the PS region close to the chemical interface defined by the two neighboring PS and PMMA nanodomains.…”

Protein Nanopatterning on Self-Organized Poly(styrene-b-isoprene) Thin Film Templates

“…20 (However, there are a variety of physical and chemical factors that can influence protein adsorption beyond the hydrophilicity of the substrate.) In previously-reported work, the successful use of a hydrophobic-hydrophilic diblock copolymer system to pattern proteins employed a very low protein concentration of 4-20 µg/ml and a very short protein adsorption time of 20-60 s. [16][17][18][19] We are not aware of any reports of the formation of protein patterns at higher protein concentrations or for longer adsorption times using the copolymer templating method. In this work, a diblock copolymer with two hydrophobic components was explored to nanopattern proteins.…”

Modulating Protein Adhesion and Conformation with Block Copolymer Surfaces

“…Kumar et al reported that in a polystyrene (PS)-polymethyl methacrylate (PMMA) block copolymer, surface proteins such as IgG and protein G will preferentially absorb to the PS block [120]. Further work demonstrated that spatially arranged proteins could retain their catalytic activity [121]. Utilizing the PS-b-PMMA system, a variety of proteins including horseradish peroxidase, mushroom tyrosinase, and enhanced green fluorescent protein were patterned on the block copolymer surfaces.…”