Preparation And Characterization Of Multiple Responsive Polymer-Peptide Bioconjugate

As an emerging class of hybrid materials, stimuli-responsive materials behave like "smart" materials, which could respond quickly to adapt environment changes. Among all the stimuli-responsive materials, polymer-peptide bioconjugates have many potential advantages in biomedical and biomaterial applications as they combine the controlled thermo-sensitivity, stability, and mechanical properties of synthetic polymers with the precise chemical structure and functionality of designed bioactive peptides. Recently, peptide-guided self-assembly of synthetic polymers proved to be a versatile tool to obtain bioinspired materials with advanced structural control. Until now, very few stimuli-responsive systems that are composed of polymer-peptide bioconjugates have been developed and are responsive to environmental conditions such as temperature, pH, or ionic strength. Of these stimuli-responsive systems, most triggered self-assemblies are responsive to only one type of external condition. Obviously, the development of multi environment-sensitive triggers for self-assembly will enable more diverse biological application of polymer-peptide self-assembled materials. The present challenge, therefore, is to design and synthesis polymer-peptide conjugates with independent multi-responsiveness.Herein we put forward a novel strategy to synthesize and characterize polymer-peptide bioconjugates which are multi-response to the pH, calcium ions, and temperature. A short peptide LLEELLEELEELLEEA (LE), which tends to form a-helices in the presence of calcium chloride, was used as the peptide template here. Furthermore, several ionizable residues-glutamines are expected to endow pH sensitivity to the peptide. Due to a characteristic lower critical solution temperature (LCST) around physiological temperature, the responsive polymer of poly (N-isopropylacrylamide)(PNIPAAm) was utilized in our studies. In existing polymerization techniques, the atom transfer radical polymerization (ATRP) has been approved as one of the most powerful and easy techniques for well-defined and controllable polymerization. Therefore, ATRP strategy was chosen to synthesize PNIPAAm-LE bioconjugate. We mainly investigates PNIPAAm-LE response toward pH, calcium ions and temperature, and differences on the self-assembly behavior were compared through three aspects such as gelation property, secondary structure and thermosensitivity. The self-assembly behaviours of the peptide LE and bioconjugate PNIPAAm-LE were first investigated by hydrogel experiments. Circular dichroism (CD) spectroscopy was used to follow the folding and subsequent self-assembly of LE and PNIPAAm-LE under different conditions. Experimental results show that the self-assembly behavior of PNIPAAm-LE and LE were pH and calcium-dual responsive. For LE and PNIPAAm-LE, a transition from the random coil to the a-helix structure were detected with addition different concentration of calcium ions. However, it takes much higher concentration of Ca2+, to induce conformation changes for PNIPAAm-LE. For LE and PNIPAAm-LE, a transition from the random coil to the a-helix structure was detected when the pH value decreased from9to4. The difference between the peptide LE and PNIPAAm-LE in low pH responsiveness might be induced by the PNIPAAm chain, which interferences the repulsive interaction between Glu within the helix. Temperature-dependent turbidity experiments were then performed to confirm the influence of pH on the LCST of the bioconjugate.In conclusion, we have shown a novel and convenient route for the preparation of the bioconjugate PNIPAAm-LE utilising resin-loaded peptide as a template in the polymerization of PNIPAAm. Moreover, the conjugate exhibits multi-responsive behaviours, showing calcium ion, pH and temperature rensibilities. This study is a deeper insight on fundamental understanding of polymer-peptide hybrid materials, which are of potential utility as stimuli-responsive biomaterials.