title: Protein-resistant NTA-functionalized polymer brushes for selective and stable immobilization of histidine-tagged proteins.
abstract: Protein-resistant polymeric coatings that allow highly selective immobilization of specific biomolecules are essential for biomedical applications such as microarrays, biosensing, heterogeneous catalysis, and bioengineering. Polymer brushes are particularly interesting for this purpose because their chemical structure and physical properties can easily be tailored to meet specific needs. This article explores the functionalization of two protein-resistant polymer brushes, poly(oligoethylene glycol methacrylate) (POEGMA) and poly(hydroxyethyl methacrylate) (PHEMA), with nitrilotriacetic acid (NTA) moieties that can complex histidine-tagged (His-tagged) proteins selectively and reversibly. Using fluorescence microscopy, IR spectroscopy, X-ray photoelectron spectroscopy, surface plasmon resonanace, and ellipsometry, we demonstrate that His-tagged green fluorescent protein can be immobilized on NTA brushes with high stability and loading. The loading saturation reached for NTA-POEGMA is higher than that for NTA-PHEMA because of increased swelling of the former brush. Despite this higher loading capacity, NTA-POEGMA remained highly protein-resistant, which shows its potential for "clean" and specific protein immobilization. Finally, we showed that the preserved protein resistance of NTA-POEGMA brushes can be used to generate well-defined binary biofunctional patterns via a simple protocol of incubations and washes. These patterns may find applications in cell arraying and screening.
authors.affiliation: Melville Laboratory for Polymer Synthesis, Department of Chemistry, University of Cambridge, Lensfield Road, Cambridge CB2 1EW, UK. jeg45@cam.ac.uk
authors.forename: Julien E
authors.initials: JE
authors.lastname: Gautrot
authors.forename: Wilhelm T S
authors.initials: WT
authors.lastname: Huck
authors.forename: Martin
authors.initials: M
authors.lastname: Welch
authors.forename: Madeleine
authors.initials: M
authors.lastname: Ramstedt
mesh_terms: D015374:Biosensing Techniques / Q000379:methods; D002384:Catalysis; D006639:Histidine / Q000737:chemistry*; D055767:Immobilized Proteins / Q000737:chemistry*; D046228:Microarray Analysis / Q000379:methods; D009571:Nitrilotriacetic Acid / Q000737:chemistry*; D011092:Polyethylene Glycols / Q000737:chemistry*; D011102:Polyhydroxyethyl Methacrylate / Q000737:chemistry*; D011993:Recombinant Fusion Proteins / Q000737:chemistry*
chemical_list: D055767:Immobilized Proteins; D011993:Recombinant Fusion Proteins; C450003:poly(ethylene glycol) methacrylate polymer; D011102:Polyhydroxyethyl Methacrylate; C033223:polyhistidine; D011092:Polyethylene Glycols; D006639:Histidine; D009571:Nitrilotriacetic Acid