Structure and Dynamics Perturbations in Ubiquitin Adsorbed or Entrapped in Silica Materials Are Related to Disparate Surface Chemistries Resolved by Solid-State NMR Spectroscopy

Abstract

Protein immobilization on material surfaces is emerging as a powerful tool in the design of devices and active materials for biomedical and pharmaceutical applications as well as for catalysis. Preservation of the protein’s biological functionality is crucial to the design process and is dependent on the ability to maintain its structural and dynamical integrity while removed from the natural surroundings. The scientific techniques to validate the structure of immobilized proteins are scarce and usually provide limited information as a result of poor resolution. In this work, we benchmarked the ability of standard solid-state NMR techniques to resolve the effects of binding to dissimilar silica materials on a model protein. In particular, the interactions between ubiquitin and the surfaces of MCM41, SBA15, and silica formed in situ were tested for their influence on the structure and dynamics of the protein. It is shown that the protein’s globular fold in the free state is only slightly perturbed in the three silica materials. Local motions on a residue level that are quenched by immobilization or, conversely, that arise from the process are also detailed. NMR measurements show that these perturbations are unique to each silica material and can serve as reporters of the characteristic surface chemistry.

Lysozyme is Sterically Trapped Within the Silica Cage in Bioinspired Silica–Lysozyme Composites: A Multi-Technique Understanding of Elusive Protein–Material Interactions

“…The intensity increase of the peak in the freeze-dried composite with respect to the protein sediment can be attributed to a lower mobility of the side chain in the composite with respect to the free protein. We also note that this peak is markedly narrowed upon rehydration and, considering the high arginine content, this behavior is similar to what was observed for arginine 15 N lines in ubiquitin interacting with MCM41. , …”

“…We also note that this peak is markedly narrowed upon rehydration and, considering the high arginine content, this behavior is similar to what was observed for arginine 15 N lines in ubiquitin interacting with MCM41. 54 , 59 …”

“…We also note that this peak is markedly narrowed upon rehydration and, considering the high arginine content, this behavior is similar to what was observed for arginine 15 N lines in ubiquitin interacting with MCM41. 54,59 Solid-state NMR also offers a unique view for the characterization of the silica matrix. The 29 Si MAS NMR spectra show signals originating from siloxanes (Q 4 , around −110 ppm), single silanols (Q 3 , around −100 ppm), and geminal silanols (Q 2 , around −90 ppm), the latter being barely visible (Figure 2, refer to Scheme S1 for a summary of the observable species).…”

Activity and Stability of Nanoconfined Alpha-Amylase in Mesoporous Silica

“…In this way, we demonstrate that it is possible to obtain high-resolution 1 H NMR spectra under MAS (at 12 kHz) and low temperatures (105 K), avoiding elevated temperatures associated with fast MAS. This avoidance could be crucial for certain sensitive biological systems that might undergo conformational changes and partially/completely unfold under the conditions of fast MAS. , Note that additional signals that do not belong to α-amylase are discernible in the DNP-enhanced spectra: those from the cryoprotectant (glycerol, 1 H shifts ∼4 ppm) and the silicon polymer plug of the DNP rotor ( 1 H/ 13 C signals at 0/∼10 ppm).…”

“…This avoidance could be crucial for certain sensitive biological systems that might undergo conformational changes and partially/completely unfold under the conditions of fast MAS. 54,55 Note that additional signals that do not belong to α-amylase are discernible in the DNP-enhanced spectra: those from the cryoprotectant (glycerol, 1 H shifts ∼4 ppm) and the silicon polymer plug of the DNP rotor ( 1 H/ 13 C signals at 0/∼10 ppm). From the high enzymatic activity observed with the small CNP-G3/MSPs probe and NMR analysis, it is clear that αamylase adsorbed on MSPs is active, as illustrated in Scheme 1.…”