The peptide hormone glucagon forms amyloid fibrils with two coexisting β-strand conformations

Gelenter, Martin D.; Smith, Katelyn J.; Liao, Shu-Yu; Mandala, Venkata S.; Dregni, Aurelio J.; Lamm, Matthew S.; Tian, Yu; Xu, Wei; Pochan, Darrin J.; Tucker, Thomas J.; Su, Yongchao; Hong, Mei

doi:10.1038/s41594-019-0238-6

Cited by 68 publications

(95 citation statements)

References 62 publications

(77 reference statements)

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“…To investigate the possible structures that are consistent with these experimental chemical shifts and long-range contacts, we conducted structure calculations for residues 268 to 340 in XPLOR-NIH (40), using chemical-shift restrained torsion angles and the sparse long-range contacts as input parameters. We assume parallel-in-register intermolecular packing, which is found in most amyloid fibrils to date (41). The lowest-energy structural model is shown in Fig.…”

Section: N4r Tau Forms a Single Ultrastructural Fibril Morphology Inmentioning

confidence: 99%

In vitro 0N4R tau fibrils contain a monomorphic β-sheet core enclosed by dynamically heterogeneous fuzzy coat segments

Dregni

Mandala

et al. 2019

Proc. Natl. Acad. Sci. U.S.A.

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212

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Misfolding of the microtubule-binding protein tau into filamentous aggregates is characteristic of many neurodegenerative diseases such as Alzheimer’s disease and progressive supranuclear palsy. Determining the structures and dynamics of these tau fibrils is important for designing inhibitors against tau aggregation. Tau fibrils obtained from patient brains have been found by cryo-electron microscopy to adopt disease-specific molecular conformations. However, in vitro heparin-fibrillized 2N4R tau, which contains all four microtubule-binding repeats (4R), was recently found to adopt polymorphic structures. Here we use solid-state NMR spectroscopy to investigate the global fold and dynamics of heparin-fibrillized 0N4R tau. A single set of 13C and 15N chemical shifts was observed for residues in the four repeats, indicating a single β-sheet conformation for the fibril core. This rigid core spans the R2 and R3 repeats and adopts a hairpin-like fold that has similarities to but also clear differences from any of the polymorphic 2N4R folds. Obtaining a homogeneous fibril sample required careful purification of the protein and removal of any proteolytic fragments. A variety of experiments and polarization transfer from water and mobile side chains indicate that 0N4R tau fibrils exhibit heterogeneous dynamics: Outside the rigid R2–R3 core, the R1 and R4 repeats are semirigid even though they exhibit β-strand character and the proline-rich domains undergo large-amplitude anisotropic motions, whereas the two termini are nearly isotropically flexible. These results have significant implications for the structure and dynamics of 4R tau fibrils in vivo.

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Section: N4r Tau Forms a Single Ultrastructural Fibril Morphology Inmentioning

confidence: 99%

In vitro 0N4R tau fibrils contain a monomorphic β-sheet core enclosed by dynamically heterogeneous fuzzy coat segments

Dregni

Mandala

et al. 2019

Proc. Natl. Acad. Sci. U.S.A.

Self Cite

212

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“…The fibrils of this protein also present a high pH dependence with respect to stability [ 155 ]. Proteins such as the Hen-egg lysozyme [ 156 ], the regulatory ATPase variant A (RavA) from E.coli [ 157 ], TTR [ 158 ], serum amyloid A [ 159 ], insulin [ 160 ], and glucagon [ 161 ], among others, have been shown to aggregate at acidic conditions (pH = 2–5), where the transition to a β-sheet structure upon oligomerization and amyloid aggregation was observed. However, other peptides and proteins, such as the designed EASZ model peptide [ 162 ], Ac-Phe-Phe-Cys-NH 2 (Ac-FFC-NH 2 ) [ 61 ] amyloid peptide model, and some peptides from human Pbx-regulating protein-1, have been shown to aggregate to β-sheet structure at basic pHs [ 163 ].…”

Section: Circular Dichroism Makes It Possible To Follow Secondary mentioning

confidence: 99%

Evaluation of Peptide/Protein Self-Assembly and Aggregation by Spectroscopic Methods

2020

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The self-assembly of proteins is an essential process for a variety of cellular functions including cell respiration, mobility and division. On the other hand, protein or peptide misfolding and aggregation is related to the development of Parkinson’s disease and Alzheimer’s disease, among other aggregopathies. As a consequence, significant research efforts are directed towards the understanding of this process. In this review, we are focused on the use of UV-Visible Absorption Spectroscopy, Fluorescence Spectroscopy and Circular Dichroism to evaluate the self-organization of proteins and peptides in solution. These spectroscopic techniques are commonly available in most chemistry and biochemistry research laboratories, and together they are a powerful approach for initial as well as routine evaluation of protein and peptide self-assembly and aggregation under different environmental stimulus. Furthermore, these spectroscopic techniques are even suitable for studying complex systems like those in the food industry or pharmaceutical formulations, providing an overall idea of the folding, self-assembly, and aggregation processes, which is challenging to obtain with high-resolution methods. Here, we compiled and discussed selected examples, together with our results and those that helped us better to understand the process of protein and peptide aggregation. We put particular emphasis on the basic description of the methods as well as on the experimental considerations needed to obtain meaningful information, to help those who are just getting into this exciting area of research. Moreover, this review is particularly useful to those out of the field who would like to improve reproducibility in their cellular and biomedical experiments, especially while working with peptide and protein systems as an external stimulus. Our final aim is to show the power of these low-resolution techniques to improve our understanding of the self-assembly of peptides and proteins and translate this fundamental knowledge in biomedical research or food applications.

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“…This is likely due to a variety of distinct but simultaneous intermediate steps occurring within the same aggregation reaction 15 . Differences in morphology often reflect different molecular arrangements and physicalchemical features [16][17][18][19][20][21][22] . Variations in 1) the number of protofilaments constituting the mature fibril, 2) the relative arrangement of the protofilaments and 3) the molecular structure of the protofilament are reported as the main sources of such variability 23,24 .…”

Section: Introductionmentioning

confidence: 99%

Probing ensemble polymorphism and single aggregate structural heterogeneity in insulin amyloid self-assembly

Luca

Galparsoro

Sancataldo

et al. 2020

Journal of Colloid and Interface Science

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Ensembles of protein aggregates are characterized by a nano-and micro-scale heterogeneity of the species. This diversity translates into a variety of effects that protein aggregates may have in biological systems, both in connection to neurodegenerative diseases and immunogenic risk of protein drug products. Moreover, this naturally occurring variety offers unique opportunities in the field of protein-based biomaterials. In the above-mentioned fields, the isolation and structural analysis of the different amyloid types within the same ensemble remain a priority, still representing a significant experimental challenge. Here we address such complexity in the case of insulin for its relevance as biopharmaceutical and its involvement in insulin-derived amyloidosis. By combining Fourier Transform Infrared Microscopy (micro-FTIR) and fluorescence lifetime imaging microscopy (FLIM) we show the occurrence, within the same ensemble of insulin protein aggregates, of a variable -structure architecture and content not only dependent on the species analyzed (spherulites or fibrils), but also on the position within a single spherulite at submicron scale. We unambiguously reveal that the surface of the spherulites are characterized by -structures with an enhanced H-bond coupling compared to the core. This information, inaccessible via bulk methods, allows us to relate the aggregate structure at molecular level to the overall morphology of the aggregates. Our findings robustly solve the problem of probing the ensemble and single particle heterogeneity of amyloid samples. Furthermore, they offer a unique, scalable and ready-to-use screening methodology for in-depth characterization of self-assembled structures, being this translatable to material sciences, drug quality control and clinical imaging of amyloid-affected tissues.

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The peptide hormone glucagon forms amyloid fibrils with two coexisting β-strand conformations

Cited by 68 publications

References 62 publications

In vitro 0N4R tau fibrils contain a monomorphic β-sheet core enclosed by dynamically heterogeneous fuzzy coat segments

In vitro 0N4R tau fibrils contain a monomorphic β-sheet core enclosed by dynamically heterogeneous fuzzy coat segments

Evaluation of Peptide/Protein Self-Assembly and Aggregation by Spectroscopic Methods

Probing ensemble polymorphism and single aggregate structural heterogeneity in insulin amyloid self-assembly

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