Integrating cryo-EM and NMR data

Geraets, James A.; Pothula, Karunakar R.; Schröder, Gunnar F.

doi:10.1016/j.sbi.2020.01.008

Cited by 25 publications

(25 citation statements)

References 48 publications

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“…In contrast to X-ray crystallography and cryo-EM, solution nuclear magnetic resonance (NMR) spectroscopy is highly suited for investigating IDPs/IDRs at amino acid-level resolution ( Gibbs et al., 2017 ; Brutscher et al., 2015 ). Solution NMR provides an equilibrium averaged readout that does not require conformational homogeneity and readily resolves conformationally dynamic regions.…”

Section: Structural Characterization Of Idps/idrsmentioning

confidence: 99%

“…Thus, NMR is amenable to investigating proteins that sample a broad conformational space. Various experiments allow for investigation of structural properties, including the identification of transient secondary structure and assessment of the magnitude and timescale of conformational dynamics ( Gibbs et al., 2017 ; Jensen et al., 2014 ). In addition, the effects of PTMs and mechanisms of binding can straightforwardly be studied ( Theillet et al., 2012 ).…”

Section: Structural Characterization Of Idps/idrsmentioning

confidence: 99%

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Characterization of functional disordered regions within chromatin-associated proteins

Musselman

Kutateladze

2021

iScience

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Summary Intrinsically disordered regions (IDRs) are abundant and play important roles in the function of chromatin-associated proteins (CAPs). These regions are often found at the N- and C-termini of CAPs and between structured domains, where they can act as more than just linkers, directly contributing to function. IDRs have been shown to contribute to substrate binding, act as auto-regulatory regions, and drive liquid-liquid droplet formation. Their disordered nature provides increased functional diversity and allows them to be easily regulated through post-translational modification. However, these regions can be especially challenging to characterize on a structural level. Here, we review the prevalence of IDRs in CAPs, highlighting several studies that address their importance in CAP function and show progress in structural characterization of these regions. A focus is placed on the unique opportunity to apply nuclear magnetic resonance (NMR) spectroscopy alongside cryo-electron microscopy to characterize IDRs in CAPs.

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Section: Structural Characterization Of Idps/idrsmentioning

confidence: 99%

Section: Structural Characterization Of Idps/idrsmentioning

confidence: 99%

Characterization of functional disordered regions within chromatin-associated proteins

Musselman

Kutateladze

2021

iScience

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“…In the 60 years since the first atomic structure of the protein myoglobin was solved using X-ray diffraction of protein crystals (Kendrew et al, 1960), the field of structural biology has been dominated by the study of globular proteins with a well-defined tertiary structure, with more than 160,000 unique structures solved to date using crystallography, NMR or cryoEM (Geraets et al, 2020). Despite this feat, more than 50% of the proteins in eukaryotes are now known to have at least one long (>30 residues) sequence that is intrinsically disordered [intrinsically disordered regions (IDRs)] and also 12% of eukaryotic proteins are completely intrinsically disordered [intrinsically disordered proteins (IDPs)] (Dunker et al, 2000;Ward et al, 2004;Tompa, 2009).…”

Section: Introductionmentioning

confidence: 99%

Looking Beyond the Core: The Role of Flanking Regions in the Aggregation of Amyloidogenic Peptides and Proteins

2020

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Amyloid proteins are involved in many neurodegenerative disorders such as Alzheimer’s disease [Tau, Amyloid β (Aβ)], Parkinson’s disease [alpha-synuclein (αSyn)], and amyotrophic lateral sclerosis (TDP-43). Driven by the early observation of the presence of ordered structure within amyloid fibrils and the potential to develop inhibitors of their formation, a major goal of the amyloid field has been to elucidate the structure of the amyloid fold at atomic resolution. This has now been achieved for a wide variety of sequences using solid-state NMR, microcrystallography, X-ray fiber diffraction and cryo-electron microscopy. These studies, together with in silico methods able to predict aggregation-prone regions (APRs) in protein sequences, have provided a wealth of information about the ordered fibril cores that comprise the amyloid fold. Structural and kinetic analyses have also shown that amyloidogenic proteins often contain less well-ordered sequences outside of the amyloid core (termed here as flanking regions) that modulate function, toxicity and/or aggregation rates. These flanking regions, which often form a dynamically disordered “fuzzy coat” around the fibril core, have been shown to play key parts in the physiological roles of functional amyloids, including the binding of RNA and in phase separation. They are also the mediators of chaperone binding and membrane binding/disruption in toxic amyloid assemblies. Here, we review the role of flanking regions in different proteins spanning both functional amyloid and amyloid in disease, in the context of their role in aggregation, toxicity and cellular (dys)function. Understanding the properties of these regions could provide new opportunities to target disease-related aggregation without disturbing critical biological functions.

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“…Cryo-EM is another complementary technique traditionally used to determine the tertiary structures of large protein complexes [ 403 , 404 ]. Unlike SAXS and NMR, samples are frozen in a thin layer of vitrified ice and density maps generated from the scattering data were obtained for a large number of individual particles, thereby producing a 3D envelope for an ensemble average of particles [ 405 ]. A particular advantage of cryo-EM is that densities for individual molecules that exist in different conformations or oligomerization states can be “binned,” enabling studies of samples containing structurally heterogeneous molecules.…”

Section: Nmr-based Hybrid Approachesmentioning

confidence: 99%

NMR Studies of Retroviral Genome Packaging

Boyd

Brown

et al. 2020

Viruses

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Nearly all retroviruses selectively package two copies of their unspliced RNA genomes from a cellular milieu that contains a substantial excess of non-viral and spliced viral RNAs. Over the past four decades, combinations of genetic experiments, phylogenetic analyses, nucleotide accessibility mapping, in silico RNA structure predictions, and biophysical experiments were employed to understand how retroviral genomes are selected for packaging. Genetic studies provided early clues regarding the protein and RNA elements required for packaging, and nucleotide accessibility mapping experiments provided insights into the secondary structures of functionally important elements in the genome. Three-dimensional structural determinants of packaging were primarily derived by nuclear magnetic resonance (NMR) spectroscopy. A key advantage of NMR, relative to other methods for determining biomolecular structure (such as X-ray crystallography), is that it is well suited for studies of conformationally dynamic and heterogeneous systems—a hallmark of the retrovirus packaging machinery. Here, we review advances in understanding of the structures, dynamics, and interactions of the proteins and RNA elements involved in retroviral genome selection and packaging that are facilitated by NMR.

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Integrating cryo-EM and NMR data

Cited by 25 publications

References 48 publications

Characterization of functional disordered regions within chromatin-associated proteins

Characterization of functional disordered regions within chromatin-associated proteins

Looking Beyond the Core: The Role of Flanking Regions in the Aggregation of Amyloidogenic Peptides and Proteins

NMR Studies of Retroviral Genome Packaging

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