title: Simple surface functionalization of polymersomes using non-antibacterial peptide anchors.
abstract: BACKGROUND
  Hollow vesicles formed from block copolymers, so-called polymersomes, have been extensively studied in the last decade for their various applications in drug delivery, in diagnostics and as nanoreactors. The immobilization of proteins on the polymersomes' surface can aid in cell targeting, lead to functional biosensors or add an additional reaction space for multistep syntheses. In almost all surface functionalization strategies to date, a chemical pre-conjugation of the polymer with a reactive group or ligand and the functionalization of the protein are required. To avoid chemical pre-conjugation, we investigated the simple and quick functionalization of preformed poly(2-methyloxazoline)-poly(dimethylsiloxane)-poly(2-methyloxazoline) (PMOXA-PDMS-PMOXA) polymersomes through the spontaneous insertion of four hydrophobic, non-antibacterial peptide anchors into the membrane to display enhanced green fluorescent protein (eGFP) on the polymersomes' surface.
  
  
  RESULTS
  Three of the four hydrophobic peptides, the transmembrane domains of a eukaryotic cytochrome b 5 , of the viral lysis protein L and of the yeast syntaxin VAM3 could be recombinantly expressed as soluble eGFP-fusion proteins and spontaneously inserted into the polymeric membrane. Characterization of the surface functionalization revealed that peptide insertion was linearly dependent on the protein concentration and possible at a broad temperature range of 4-42 °C. Up to 2320 ± 280 eGFP molecules were immobilized on a single polymersome, which is in agreement with the calculated maximum loading capacity. The peptide insertion was stable without disrupting membrane integrity as shown in calcein leakage experiments and the functionalized polymersomes remained stable for at least 6 weeks.
  
  
  CONCLUSIONS
  The surface functionalization of polymersomes with hydrophilic proteins can be mediated by several peptide anchors in a spontaneous process at extremely mild insertion conditions and without the need of pre-conjugating polymers.
authors.affiliation: Institute of Biochemical Engineering, Technical University of Munich, Boltzmannstraße 15, 85748, Garching, Germany.
authors.forename: Ludwig
authors.initials: L
authors.lastname: Klermund
authors.affiliation: Institute of Biochemical Engineering, Technical University of Munich, Boltzmannstraße 15, 85748, Garching, Germany.
authors.forename: Sarah T
authors.initials: ST
authors.lastname: Poschenrieder
authors.affiliation: Institute of Biochemical Engineering, Technical University of Munich, Boltzmannstraße 15, 85748, Garching, Germany. k.castiglione@lrz.tum.de.
authors.forename: Kathrin
authors.initials: K
authors.lastname: Castiglione
mesh_terms: D003001:Cloning, Molecular / Q000379:methods; D004337:Drug Carriers / Q000737:chemistry; D016503:Drug Delivery Systems; D004926:Escherichia coli / Q000235:genetics; D049452:Green Fluorescent Proteins / Q000737:chemistry* / Q000235:genetics; D057927:Hydrophobic and Hydrophilic Interactions; D055767:Immobilized Proteins / Q000737:chemistry* / Q000235:genetics; D018613:Microscopy, Confocal; D008856:Microscopy, Fluorescence; D010080:Oxazoles / Q000737:chemistry*; D010455:Peptides / Q000737:chemistry* / Q000235:genetics; D011108:Polymers / Q000737:chemistry*; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics; D013499:Surface Properties
chemical_list: D004337:Drug Carriers; D055767:Immobilized Proteins; D010080:Oxazoles; D010455:Peptides; D011108:Polymers; D011993:Recombinant Fusion Proteins; C485184:enhanced green fluorescent protein; C542789:poly(2-methyl oxazoline)-block polydimethylsiloxane-block-poly(2-methyl oxazoline); D049452:Green Fluorescent Proteins
keywords: Block copolymer; Hydrophobic peptide anchor; Non-antibacterial peptide; PMOXA-PDMS-PMOXA; Polymersomes; Surface functionalization