ABSTRACT
We present a facile protocol for the controlled growth of highly oriented and polyoxometalate-incorporating HKUST-1 SURMOFs. Combining the spin-coating technique with alcohol-vapour induced growth, film thickness, crystallite orientation and crystal size can be precisely tuned. The SURMOFs exhibit excellent abilities in selective adsorption of cationic dyes and water oxidation.
ABSTRACT
Transmission electron cryo-microscopy (cryoEM) of vitrified biological specimens is a powerful tool for structural biology. Current preparation of vitrified biological samples starts off with sample isolation and purification, followed by the fixation in a freestanding layer of amorphous ice. Here, we demonstrate that ultrathin (â¼10 nm) smart molecular nanosheets having specific biorecognition sites embedded in a biorepulsive layer covalently bound to a mechanically stable carbon nanomembrane allow for a much simpler isolation and structural analysis. We characterize in detail the engineering of these nanosheets and their biorecognition properties employing complementary methods such as X-ray photoelectron and infrared spectroscopy, atomic force microscopy as well as surface plasmon resonance measurements. The desired functionality of the developed nanosheets is demonstrated by in situ selection of a His-tagged protein from a mixture and its subsequent structural analysis by cryoEM.
Subject(s)
Carbon , Electrons , Cryoelectron Microscopy , Microscopy, Atomic Force , Microscopy, Electron, TransmissionABSTRACT
In electron cryo-microscopy, structure determination of protein molecules is frequently hampered by adsorption of the particles to the support film material, typically amorphous carbon. Here, we report that pyrene derivatives with one or two polyglycerol (PG) side chains bind to the amorphous carbon films, forming a biorepulsive hydrogel layer so that the number of protein particles in the vitreous ice drastically increases. This approach could be extended by adding a hydrogel-functionalized carbon nanotube network (HyCaNet, the hydrogel again being formed from the PG-pyrene derivatives), which stabilized the protein-containing thin ice films during imaging with the electron beam. The stabilization resulted in reduced particle motion by up to 70%. These substrates were instrumental for determining the structure of a large membrane protein complex.
Subject(s)
Cryoelectron Microscopy/methods , Hydrogels/chemistry , Membrane Proteins/chemistry , Detergents/chemistry , Glycerol/chemistry , Membrane Proteins/ultrastructure , Nanotubes/chemistry , Polymers/chemistry , Protein Stability , Pyrenes/chemistry , VitrificationABSTRACT
We developed a method to improve specimen preparation for electron cryo-microscopy of membrane proteins. The method features a perforated hydrogel nanomembrane that stabilizes the thin film of aqueous buffer spanning the holes of holey carbon films, while at the same time preventing the depletion of protein molecules from these holes. The membrane is obtained by cross-linking of thiolated polyglycerol dendrimer films on gold, which self-perforate upon transfer to holey carbon substrates, forming a sub-micron-sized hydrogel network. The perforated nanomembrane improves the distribution of the protein molecules in the ice considerably. This facilitates data acquisition as demonstrated with two eukaryotic membrane protein complexes.
ABSTRACT
As well-oriented, surface-bound metal-organic frameworks become the centerpiece of many new applications, a profound understanding of their growth mode becomes necessary. This work shows that the currently favored model of surface templating is in fact a special case valid only for systems with a more or less cubic crystal shape, while in less symmetric systems crystal ripening and minimization of surface energies dominate the growth process.
