TY - JOUR A1 - Charan, Himanshu A1 - Kinzel, Julia A1 - Glebe, Ulrich A1 - Anand, Deepak A1 - Garakani, Tayebeh Mirzaei A1 - Zhu, Leilei A1 - Bocola, Marco A1 - Schwaneberg, Ulrich A1 - Böker, Alexander T1 - Grafting PNIPAAm from beta-barrel shaped transmembrane nanopores JF - Biomaterials : biomaterials reviews online N2 - The research on protein-polymer conjugates by grafting from the surface of proteins has gained significant interest in the last decade. While there are many studies with globular proteins, membrane proteins have remained untouched to the best of our knowledge. In this study, we established the conjugate formation with a class of transmembrane proteins and grow polymer chains from the ferric hydroxamate uptake protein component A (FhuA; a beta-barrel transmembrane protein of Escherichia coli). As the lysine residues of naturally occurring FhuA are distributed over the whole protein, FhuA was reengineered to have up to 11 lysines, distributed symmetrically in a rim on the membrane exposed side (outside) of the protein channel and exclusively above the hydrophobic region. Reengineering of FhuA ensures a polymer growth only on the outside of the beta-barrel and prevents blockage of the channel as a result of the polymerization. A water-soluble initiator for controlled radical polymerization (CRP) was consecutively linked to the lysine residues of FhuA and N-isopropylacrylamide (NIPAAm) polymerized under copper mediated CRP conditions. The conjugate formation was analyzed by using MALDI-ToF mass spectrometry, SDS-PAGE, circular dichroism spectroscopy, analytical ultracentrifugation, dynamic light scattering, transmission electron microscopy and size exclusion chromatography. Such conjugates combine the specific functions of the transmembrane proteins, like maintaining membrane potential gradients or translocation of substrates with the unique properties of synthetic polymers such as temperature and pH stimuli handles. FhuA-PNIPAAm conjugates will serve as functional nanosized building blocks for applications in targeted drug delivery, self-assembly systems, functional membranes and transmembrane protein gated nanoreactors. (C) 2016 Elsevier Ltd. All rights reserved. KW - Transmembrane protein KW - FhuA KW - Protein-polymer conjugate KW - Grafting-from polymerization KW - NIPAAm KW - BBTP Y1 - 2016 U6 - https://doi.org/10.1016/j.biomaterials.2016.08.033 SN - 0142-9612 SN - 1878-5905 VL - 107 SP - 115 EP - 123 PB - Elsevier CY - Oxford ER - TY - JOUR A1 - Charan, Himanshu A1 - Glebe, Ulrich A1 - Anand, Deepak A1 - Kinzel, Julia A1 - Zhu, Leilei A1 - Bocola, Marco A1 - Garakani, Tayebeh Mirzaei A1 - Schwaneberg, Ulrich A1 - Böker, Alexander T1 - Nano-thin walled micro-compartments from transmembrane protein-polymer conjugates JF - Soft matter N2 - The high interfacial activity of protein-polymer conjugates has inspired their use as stabilizers for Pickering emulsions, resulting in many interesting applications such as synthesis of templated micro-compartments and protocells or vehicles for drug and gene delivery. In this study we report, for the first time, the stabilization of Pickering emulsions with conjugates of a genetically modified transmembrane protein, ferric hydroxamate uptake protein component A (FhuA). The lysine residues of FhuA with open pore (FhuA Delta CVFtev) were modified to attach an initiator and consequently controlled radical polymerization (CRP) carried out via the grafting-from technique. The resulting conjugates of FhuA Delta CVFtev with poly(N-isopropylacrylamide) (PNIPAAm) and poly((2-dimethylamino) ethyl methacrylate) (PDMAEMA), the so-called building blocks based on transmembrane proteins (BBTP), have been shown to engender larger structures. The properties such as pH-responsivity, temperature-responsivity and interfacial activity of the BBTP were analyzed using UV-Vis spectrophotometry and pendant drop tensiometry. The BBTP were then utilized for the synthesis of highly stable Pickering emulsions, which could remain non-coalesced for well over a month. A new UV-crosslinkable monomer was synthesized and copolymerized with NIPAAm from the protein. The emulsion droplets, upon crosslinking of polymer chains, yielded micro-compartments. Fluorescence microscopy proved that these compartments are of micrometer scale, while cryo-scanning electron microscopy and scanning force microscopy analysis yielded a thickness in the range of 11.1 +/- 0.6 to 38.0 +/- 18.2 nm for the stabilizing layer of the conjugates. Such micro-compartments would prove to be beneficial in drug delivery applications, owing to the possibility of using the channel of the transmembrane protein as a gate and the smart polymer chains as trigger switches to tune the behavior of the capsules. Y1 - 2017 U6 - https://doi.org/10.1039/c6sm02520j SN - 1744-683X SN - 1744-6848 VL - 13 SP - 2866 EP - 2875 PB - Royal Society of Chemistry CY - Cambridge ER -