9 results on '"Griffin, Robert G."'
Search Results
2. Structural Characterization of E22G Aβ1-42 Fibrils via 1H detected MAS NMR
- Author
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Golota, Natalie, primary, Michael, Brian, additional, Saliba, Edward P., additional, Linse, Sara, additional, and Griffin, Robert G., additional
- Published
- 2024
- Full Text
- View/download PDF
3. Design of Zirconia Mas Rotors for Dynamic Nuclear Polarization Nmr
- Author
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Li, Guangjiang, primary, Dastrup, Blake, additional, Palani, Ravi Shankar, additional, Shapiro, Michael A., additional, Jawla, Sudheer K., additional, Griffin, Robert G., additional, and Temkin, Richard J., additional
- Published
- 2024
- Full Text
- View/download PDF
4. Structural characterization of E22G Aβ1–42 fibrils via1H detected MAS NMR.
- Author
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Golota, Natalie C., Michael, Brian, Saliba, Edward P., Linse, Sara, and Griffin, Robert G.
- Abstract
Amyloid fibrils have been implicated in the pathogenesis of several neurodegenerative diseases, the most prevalent example being Alzheimer's disease (AD). Despite the prevalence of AD, relatively little is known about the structure of the associated amyloid fibrils. This has motivated our studies of fibril structures, extended here to the familial Arctic mutant of Aβ
1–42 , E22G-Aβ1–42 . We found E22G-AβM0,1–42 is toxic to Escherichia coli, thus we expressed E22G-Aβ1–42 fused to the self-cleavable tag NPro in the form of its EDDIE mutant. Since the high surface activity of E22G-Aβ1–42 makes it difficult to obtain more than sparse quantities of fibrils, we employed1 H detected magic angle spinning (MAS) nuclear magnetic resonance (NMR) experiments to characterize the protein. The1 H detected13 C–13 C methods were first validated by application to fully protonated amyloidogenic nanocrystals of GNNQQNY, and then applied to fibrils of the Arctic mutant of Aβ, E22G-Aβ1–42 . The MAS NMR spectra indicate that the biosynthetic samples of E22G-Aβ1–42 fibrils comprise a single conformation with13 C chemical shifts extracted from hCH, hNH, and hCCH spectra that are very similar to those of wild type Aβ1–42 fibrils. These results suggest that E22G-Aβ1–42 fibrils have a structure similar to that of wild type Aβ1–42 . [ABSTRACT FROM AUTHOR]- Published
- 2024
- Full Text
- View/download PDF
5. Lipid Dynamics and Protein–Lipid Interactions in 2D Crystals Formed with the β-Barrel Integral Membrane Protein VDAC1
- Author
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Eddy, Matthew T., Ong, Ta-Chung, Clark, Lindsay, Teijido, Oscar, van der Wel, Patrick C. A., Garces, Robert, Wagner, Gerhard, Rostovtseva, Tatiana K., and Griffin, Robert G.
- Abstract
We employ a combination of 13C/15N magic angle spinning (MAS) NMR and 2H NMR to study the structural and functional consequences of different membrane environments on VDAC1 and, conversely, the effect of VDAC1 on the structure of the lipid bilayer. MAS spectra reveal a well-structured VDAC1 in 2D crystals of dimyristoylphosphatidylcholine (DMPC) and diphytanoylphosphatidylcholine (DPhPC), and their temperature dependence suggests that the VDAC structure does not change conformation above and below the lipid phase transition temperature. The same data show that the N-terminus remains structured at both low and high temperatures. Importantly, functional studies based on electrophysiological measurements on these same samples show fully functional channels, even without the presence of Triton X-100 that has been found necessary for in vitro-refolded channels. 2H solid-state NMR and differential scanning calorimetry were used to investigate the dynamics and phase behavior of the lipids within the VDAC1 2D crystals. 2H NMR spectra indicate that the presence of protein in DMPC results in a broad lipid phase transition that is shifted from 19 to ∼27 °C and show the existence of different lipid populations, consistent with the presence of both annular and bulk lipids in the functionally and structurally homogeneous samples.
- Published
- 2024
- Full Text
- View/download PDF
6. Proton-Assisted Recoupling (PAR) in Peptides and Proteins
- Author
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Donovan, Kevin J., Jain, Sheetal K., Silvers, Robert, Linse, Sara, and Griffin, Robert G.
- Abstract
Proton-assisted recoupling (PAR) is examined by exploring optimal experimental conditions and magnetization transfer rates in a variety of biologically relevant nuclear spin-systems, including simple amino acids, model peptides, and two proteins–nanocrystalline protein G (GB1), and importantly amyloid beta 1–42 (M0Aβ1–42) fibrils. A selective PAR protocol, SUBPAR (setting up better proton assisted recoupling), is described to observe magnetization transfer in one-dimensional spectra, which minimizes experiment time (in comparison to two-dimensional experiments) and thereby enables an efficient assessment of optimal PAR conditions for a desired magnetization transfer. In the case of the peptide spin systems, experimental and simulated PAR data sets are compared on a semiquantitative level, thereby elucidating the interactions influencing PAR magnetization transfer and their manifestations in different spin transfer networks. Using the optimum Rabi frequencies determined by SUBPAR, PAR magnetization transfer trajectories (or buildup curves) were recorded and compared to simulated results for short peptides. PAR buildup curves were also recorded for M0Aβ1–42and examined conjointly with a recent structural model. The majority of salient cross-peak intensities observed in the M0Aβ1–42PAR spectra are well-modeled with a simple biexponential equation, although the fitting parameters do not show any strong correlation to internuclear distances. Nevertheless, these parameters provide a wealth of invaluable semiquantitative structural constraints for the M0Aβ1–42. The results presented here offer a complete protocol for recording PAR 13C–13C correlation spectra with high-efficiency and using the resulting information in protein structural studies.
- Published
- 2024
- Full Text
- View/download PDF
7. Aggregation and Fibril Structure of AβM01–42and Aβ1–42
- Author
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Silvers, Robert, Colvin, Michael T., Frederick, Kendra K., Jacavone, Angela C., Lindquist, Susan, Linse, Sara, and Griffin, Robert G.
- Abstract
A mechanistic understanding of Aβ aggregation and high-resolution structures of Aβ fibrils and oligomers are vital to elucidating relevant details of neurodegeneration in Alzheimer’s disease, which will facilitate the rational design of diagnostic and therapeutic protocols. The most detailed and reproducible insights into structure and kinetics have been achieved using Aβ peptides produced by recombinant expression, which results in an additional methionine at the N-terminus. While the length of the C-terminus is well established to have a profound impact on the peptide’s aggregation propensity, structure, and neurotoxicity, the impact of the N-terminal methionine on the aggregation pathways and structure is unclear. For this reason, we have developed a protocol to produce recombinant Aβ1–42, sans the N-terminal methionine, using an N-terminal small ubiquitin-like modifier–Aβ1–42fusion protein in reasonable yield, with which we compared aggregation kinetics with AβM01–42containing the additional methionine residue. The data revealed that Aβ1–42and AβM01–42aggregate with similar rates and by the same mechanism, in which the generation of new aggregates is dominated by secondary nucleation of monomers on the surface of fibrils. We also recorded magic angle spinning nuclear magnetic resonance spectra that demonstrated that excellent spectral resolution is maintained with both AβM01–42and Aβ1–42and that the chemical shifts are virtually identical in dipolar recoupling experiments that provide information about rigid residues. Collectively, these results indicate that the structure of the fibril core is unaffected by N-terminal methionine. This is consistent with the recent structures of AβM01–42in which M0 is located at the terminus of a disordered 14-amino acid N-terminal tail.
- Published
- 2024
- Full Text
- View/download PDF
8. Peptidic "Molecular Beacon" for Collagen.
- Author
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Yang J, Quan Y, Ouyang Y, Tan KO, Weber RT, Griffin RG, and Raines RT
- Abstract
Collagen-mimetic peptides (CMP) have been invaluable tools for understanding the structure and function of collagen, which is the most abundant protein in animals. CMPs have also been developed as probes that detect damaged collagen because of the specificity required to form a collagen triple helix. These probes are not, however, ratiometric. Here, we used EPR spectroscopy to determine the end-to-end distances of CMPs that do not form stable homotrimeric helices. We found that those distances are shorter than the distances in the context of a collagen triple helix, suggesting their potential utility as a "molecular beacon" and guiding the choice and location of a pendant fluorophore-quencher pair. We then showed that a molecular beacon based on a glycine-(2 S ,4 S )-4-fluoroproline-(2 S ,4 R )-4-hydroxyproline tripeptide repeat and EDANS-DABCYL pair enabled the ratiometric detection of its binding to both other CMPs and natural mammalian collagen. These results provide guidance for the development of a new modality for detecting damaged collagen in physiological settings.
- Published
- 2024
- Full Text
- View/download PDF
9. Structural characterization of E22G Aβ 1-42 fibrils via 1 H detected MAS NMR.
- Author
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Golota NC, Michael B, Saliba EP, Linse S, and Griffin RG
- Subjects
- Amyloid chemistry, Amyloid metabolism, Nuclear Magnetic Resonance, Biomolecular, Escherichia coli genetics, Escherichia coli metabolism, Mutation, Humans, Amyloid beta-Peptides chemistry, Amyloid beta-Peptides genetics, Amyloid beta-Peptides metabolism, Peptide Fragments chemistry, Peptide Fragments genetics, Peptide Fragments metabolism
- Abstract
Amyloid fibrils have been implicated in the pathogenesis of several neurodegenerative diseases, the most prevalent example being Alzheimer's disease (AD). Despite the prevalence of AD, relatively little is known about the structure of the associated amyloid fibrils. This has motivated our studies of fibril structures, extended here to the familial Arctic mutant of Aβ
1-42 , E22G-Aβ1-42 . We found E22G-AβM0,1-42 is toxic to Escherichia coli , thus we expressed E22G-Aβ1-42 fused to the self-cleavable tag NPro in the form of its EDDIE mutant. Since the high surface activity of E22G-Aβ1-42 makes it difficult to obtain more than sparse quantities of fibrils, we employed1 H detected magic angle spinning (MAS) nuclear magnetic resonance (NMR) experiments to characterize the protein. The1 H detected13 C-13 C methods were first validated by application to fully protonated amyloidogenic nanocrystals of GNNQQNY, and then applied to fibrils of the Arctic mutant of Aβ, E22G-Aβ1-42 . The MAS NMR spectra indicate that the biosynthetic samples of E22G-Aβ1-42 fibrils comprise a single conformation with13 C chemical shifts extracted from hCH, hNH, and hCCH spectra that are very similar to those of wild type Aβ1-42 fibrils. These results suggest that E22G-Aβ1-42 fibrils have a structure similar to that of wild type Aβ1-42 .- Published
- 2024
- Full Text
- View/download PDF
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