Temperature-Controlled Orientation of Proteins on Temperature-Responsive Grafted Polymer Brushes: Poly(butyl methacrylate) vs Poly(butyl acrylate): Morphology, Wetting, and Protein Adsorption

Abstract

Poly(n-butyl methacrylate) (PBMA) or poly(n-butyl acrylate) (PBA)-grafted brush coatings attached to glass were successfully prepared using atom-transfer radical polymerization "from the surface". The thicknesses and composition of the PBMA and PBA coatings were examined using ellipsometry and time-of-flight secondary ion mass spectrometry (ToF-SIMS), respectively. For PBMA, the glasstransition temperature constitutes a range close to the physiological limit, which is in contrast to PBA, where the glass-transition temperature is around −55 °C. Atomic force microscopy studies at different temperatures suggest a strong morphological transformation for PBMA coatings, in contrast to PBA, where such essential changes in the surface morphology are absent. Besides, for PBMA coatings, protein adsorption depicts a strong temperature dependence. The combination of bovine serum albumin and anti-IgG structure analysis with the principal component analysis of ToF-SIMS spectra revealed a different orientation of proteins adsorbed to PBMA coatings at different temperatures. In addition, the biological activity of anti-IgG molecules adsorbed at different temperatures was evaluated through tracing the specific binding with goat IgG.

Controlling orientation, conformation, and biorecognition of proteins on silane monolayers, conjugate polymers, and thermo-responsive polymer brushes: investigations using TOF-SIMS and principal component analysis

“…Therefore, the access to binding sites and antigen binging efficiency depends on antibody orientation involving flat-on (all domains attached to a surface), side-on (Fc and one Fab domain attached to a surface), head-on (both Fab domains attached to surface and Fc facing up), and tail-on (Fc domain attached to a surface and both Fab facing up) orientations [22]. TOF-SIMS with PCA was applied for the examination of the dominant orientation adopted by antibodies on a SAM modified gold [33,37,73] and silicon surface [34,74], polymer layers [28,33,35,55], and polymer brushes [75] as well as to determine the orientation of bioaffinity-bound antibodies [74,76]. The studies performed so far have provided a comparison of a dominant antibody orientation on different surfaces and a definition of the factors determining protein orientation, the being latter discussed in detail in "Factors affecting protein orientation: examination illustrated with the antibody on self-assembled monolayers" of this review.…”

“…One solution to this problem is a simultaneous TOF-SIMS examination of reference samples with immobilized Fc and Fab domains [33,34,73]. Recently, Awsiuk et al proposed to interpret the orientational order of an examined IgG antibody with an unknown amino acid composition of its domains, using the relation between the just obtained PCA loadings and the PCA loadings determined earlier [73] for an IgG with a known complete amino acid composition of the Fab and Fc domains [35,75]. This idea is based on the fact that the relative prevalence of amino acids contributing to each signal and originating from F(ab') 2 versus Fc fragment is largely preserved for different antibodies regardless of their exact composition (as shown in [74] for the amino acid composition ratio in F(ab') 2 and Fc subunits).…”

“…Relations between orientation and electrostatic interactions as well as biorecognition efficiency were reported. So far, the TOF-SIMS technique has been successfully applied to analyze the surface orientation of proteins such as bovine serum albumin (BSA) [75], fibronectin [79], lysozyme [80], osteocalcin [81], cytochrome b5 [82], and β-lactoglobuline [83].…”

“…Practically all the biomedical applications of thermoresponsive polymer brushes reported so far use LCST transition as a temperature response mechanism. Recently, a novel approach to gain temperature sensitivity has been demonstrated, involving a glassy-to-rubbery state transition: in particular, examined has been a grafted brush of poly(n-butyl methacrylate) (PBMA) attached to the glass, with a glass transition temperature (T g around 13-25°C) within the range of physiological temperatures [75]. Its properties are depicted in Fig.…”

Self‐Assembled Polymeric Membranes and Nanoassemblies on Surfaces: Preparation, Characterization, and Current Applications

“…Since the membranes are covalently bonded to the surface, they possess enhanced mechanical stability and undergo degradation much slower compared to their free‐standing counterparts, which is a necessary characteristic for long‐term applications . For instance, a triblock composed of two blocks of zwitterionic sulfobetaine (SBMA) and glycidyl methacrylate grafted polymer brushes on planar surfaces also serve as functional supports to self‐assembled lipid membranes to mimic the extracellular membrane …”