Protein Micro- and Nanopatterning Using Aminosilanes with Protein-Resistant Photolabile Protecting Groups

Abstract

An approach to the integration of nanolithography with synthetic chemical methodology is described, in which near-field optical techniques are used to selectively deprotect films formed by the adsorption of aminosilanes protected by modified 2-nitrophenylethoxycarbonyl (NPEOC) groups. The NPEOC groups are functionalized at the m- or p-position with either a tetraethyleneglycol or a heptaethylene glycol adduct. We describe the synthesis of these bioresistant aminosilanes and the characterization of the resulting photoreactive films. Photodeprotection by exposure to UV light (λ = 325 nm) yielded the amine with high efficiency, at a similar rate for all four adsorbates, and was complete after an exposure of 2.24 J cm(-2). Following photodeprotection, derivatization by trifluoroacetic anhydride was carried out with high efficiency. Micropatterned samples, formed using a mask, were derivatized with aldehyde-functionalized polymer nanoparticles and, following derivatization with biotin, were used to form patterns of avidin-coated polymer particles. Fluorescence microscopy and atomic force microscopy data demonstrated that the intact protecting groups conferred excellent resistance to nonspecific adsorption. Nanometer-scale patterns were created using scanning near-field photolithography and were derivatized with biotin. Subsequent conjugation with avidin-functionalized polymer nanoparticles yielded clear fluorescence images that indicated dense attachment to the nanostructures and excellent protein resistance on the surrounding surface. These simple photocleavable protecting group strategies, combined with the use of near-field exposure, offer excellent prospects for the control of surface reactivity at nanometer resolution in biological systems and offer promise for integrating the top-down and bottom-up molecular fabrication paradigms.

Solid-State Nanostructured Materials from Self-Assembly of a Globular Protein–Polymer Diblock Copolymer

“…22–23 Alternately, a variety of lithographic methods including microcontact printing, photolithography and dip-pen lithography have been used to create templates for protein immobilization by patterning an attachment site where the protein selectively binds. 24–26 Covalent binding using lysine-NHS or cysteine-maleimide chemistries, 27 or non-covalent binding using biotin-avidin 28 or His tag-NTA 29 interactions are then employed to immobilize enzymes on the pre-formed patterns. In a third approach, nanostructured templates are self-assembled from amphiphilic molecules and the proteins are inserted into the templates.…”

Focal Molography: Coherent Microscopic Detection of Biomolecular Interaction

“…Sensitive photochemistry for the deprotection of functional groups on linker molecules by photolysis is readily available [39][40][41][42][43]. The linker in the hydrogel is connected at one end to the hydrogel, while the other terminal group on the linker is protected by a photolabile protecting group for light-induced immobilization of the receptor within the hydrogel.…”

Micrometre and nanometre scale patterning of binary polymer brushes, supported lipid bilayers and proteins

“…48 Exposure of the protected film to near-UV light leads to selective removal of the NPPOC group (Scheme 1), enabling derivatisation of free amines. 48,49 This enables spatial control of surface reactivity to be achieved at the micrometre scale using a mask, or at much shorter length scales using a near-field probe. 48,49 In the present study, we apply this strategy to the fabrication of binary polymer brushes, using a mask-based process to fabricate micrometre-scale structures and interferometric exposure to achieve high-fidelity patterning across macroscopic areas with nanometre scale resolution.…”