Molecular basis for insulin fibril assembly Ivanova, Magdalena I; Sievers, Stuart A; Sawaya, Michael R ...
Proceedings of the National Academy of Sciences - PNAS,
11/2009, Letnik:
106, Številka:
45
Journal Article
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In the rare medical condition termed injection amyloidosis, extracellular fibrils of insulin are observed. We found that the segment of the insulin B-chain with sequence LVEALYL is the smallest ...segment that both nucleates and inhibits the fibrillation of full-length insulin in a molar ratio-dependent manner, suggesting that this segment is central to the cross-β spine of the insulin fibril. In isolation from the rest of the protein, LVEALYL forms microcrystalline aggregates with fibrillar morphology, the structure of which we determined to 1 Å resolution. The LVEALYL segments are stacked into pairs of tightly interdigitated β-sheets, each pair displaying the dry steric zipper interface typical of amyloid-like fibrils. This structure leads to a model for fibrils of human insulin consistent with electron microscopic, x-ray fiber diffraction, and biochemical studies.
Many globular and natively disordered proteins can convert into amyloid fibrils. These fibrils are associated with numerous pathologies as well as with normal cellular functions, and frequently form ...during protein denaturation. Inhibitors of pathological amyloid fibril formation could be useful in the development of therapeutics, provided that the inhibitors were specific enough to avoid interfering with normal processes. Here we show that computer-aided, structure-based design can yield highly specific peptide inhibitors of amyloid formation. Using known atomic structures of segments of amyloid fibrils as templates, we have designed and characterized an all-D-amino-acid inhibitor of the fibril formation of the tau protein associated with Alzheimer's disease, and a non-natural L-amino-acid inhibitor of an amyloid fibril that enhances sexual transmission of human immunodeficiency virus. Our results indicate that peptides from structure-based designs can disrupt the fibril formation of full-length proteins, including those, such as tau protein, that lack fully ordered native structures. Because the inhibiting peptides have been designed on structures of dual-β-sheet 'steric zippers', the successful inhibition of amyloid fibril formation strengthens the hypothesis that amyloid spines contain steric zippers.
Towards a pharmacophore for amyloid Landau, Meytal; Sawaya, Michael R; Faull, Kym F ...
PLoS biology,
06/2011, Letnik:
9, Številka:
6
Journal Article
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Diagnosing and treating Alzheimer's and other diseases associated with amyloid fibers remains a great challenge despite intensive research. To aid in this effort, we present atomic structures of ...fiber-forming segments of proteins involved in Alzheimer's disease in complex with small molecule binders, determined by X-ray microcrystallography. The fiber-like complexes consist of pairs of β-sheets, with small molecules binding between the sheets, roughly parallel to the fiber axis. The structures suggest that apolar molecules drift along the fiber, consistent with the observation of nonspecific binding to a variety of amyloid proteins. In contrast, negatively charged orange-G binds specifically to lysine side chains of adjacent sheets. These structures provide molecular frameworks for the design of diagnostics and drugs for protein aggregation diseases.
In a strongly nonlinear discrete system, the spatial size of an excitation can become comparable to, and influenced by, the lattice spacing. Such intrinsic localized modes (ILMs)-also called ...'discrete breathers' or 'lattice solitons'-are responsible for energy localization in the dynamics of discrete nonlinear lattices. Their energy profiles resemble those of localized modes of defects in a harmonic lattice but, like solitons, they can move (although, unlike solitons, some energy is exchanged during collisions between them). The manipulation of these localized energy 'hotspots' has been achieved in systems as diverse as annular arrays of coupled Josephson junctions, optical waveguide arrays, two-dimensional nonlinear photonic crystals and micromechanical cantilever arrays. There is also some evidence for the existence of localized excitations in atomic lattices, although individual ILMs have yet to be identified. Here we report the observation of countable localized excitations in an antiferromagnetic spin lattice by means of a nonlinear spectroscopic technique. This detection capability permits the properties of individual ILMs to be probed; the disappearance of each ILM registers as a step in the time-dependent signal, with the surprising result that the energy staircase of ILM excitations is uniquely defined.
Based on the crystal structure of the cross-β spine formed by the peptide NNQQNY, we have developed a computational approach for identifying those segments of amyloidogenic proteins that themselves ...can form amyloid-like fibrils. The approach builds on experiments showing that hexapeptides are sufficient for forming amyloid-like fibrils. Each six-residue peptide of a protein of interest is mapped onto an ensemble of templates, or 3D profile, generated from the crystal structure of the peptide NNQQNY by small displacements of one of the two intermeshed β-sheets relative to the other. The energy of each mapping of a sequence to the profile is evaluated by using ROSETTADESIGN, and the lowest energy match for a given peptide to the template library is taken as the putative prediction. If the energy of the putative prediction is lower than a threshold value, a prediction of fibril formation is made. This method can reach an accuracy of ≈80% with a P value of$\supset10^{-12}$when a conservative energy threshold is used to separate peptides that form fibrils from those that do not. We see enrichment for positive predictions in a set of fibril-forming segments of amyloid proteins, and we illustrate the method with applications to proteins of interest in amyloid research.
ALS is a terminal disease of motor neurons that is characterized by accumulation of proteinaceous deposits in affected cells. Pathological deposition of mutated Cu/Zn superoxide dismutase (SOD1) ...accounts for ∼20% of the familial ALS (fALS) cases. However, understanding the molecular link between mutation and disease has been difficult, given that more than 140 different SOD1 mutants have been observed in fALS patients. In addition, the molecular origin of sporadic ALS (sALS) is unclear. By dissecting the amino acid sequence of SOD1, we identified four short segments with a high propensity for amyloid fibril formation. We find that fALS mutations in these segments do not reduce their propensity to form fibrils. The atomic structures of two fibril-forming segments from the C terminus, ¹⁰¹DSVISLS ¹⁰⁷ and ¹⁴⁷GVIGIAQ ¹⁵³, reveal tightly packed β-sheets with steric zipper interfaces characteristic of the amyloid state. Based on these structures, we conclude that both C-terminal segments are likely to form aggregates if available for interaction. Proline substitutions in ¹⁰¹DSVISLS ¹⁰⁷ and ¹⁴⁷GVIGIAQ ¹⁵³ impaired nucleation and fibril growth of full-length protein, confirming that these segments participate in aggregate formation. Our hypothesis is that improper protein maturation and incompletely folded states that render these aggregation-prone segments available for interaction offer a common molecular pathway for sALS and fALS.
Context. Millimetre-wave continuum astronomy is today an indispensable tool for both general astrophysics studies (e.g. star formation, nearby galaxies) and cosmology (e.g. cosmic microwave ...background and high-redshift galaxies). General purpose, large-field-of-view instruments are needed to map the sky at intermediate angular scales not accessible by the high-resolution interferometers (e.g. ALMA in Chile, NOEMA in the French Alps) and by the coarse angular resolution space-borne or ground-based surveys (e.g. Planck, ACT, SPT). These instruments have to be installed at the focal plane of the largest single-dish telescopes, which are placed at high altitude on selected dry observing sites. In this context, we have constructed and deployed a three-thousand-pixel dual-band (150 GHz and 260 GHz, respectively 2 mm and 1.15 mm wavelengths) camera to image an instantaneous circular field-of-view of 6.5 arcmin in diameter, and configurable to map the linear polarisation at 260 GHz. Aims. First, we are providing a detailed description of this instrument, named NIKA2 (New IRAM KID Arrays 2), in particular focussing on the cryogenics, optics, focal plane arrays based on Kinetic Inductance Detectors, and the readout electronics. The focal planes and part of the optics are cooled down to the nominal 150 mK operating temperature by means of an adhoc dilution refrigerator. Secondly, we are presenting the performance measured on the sky during the commissioning runs that took place between October 2015 and April 2017 at the 30-m IRAM telescope at Pico Veleta, near Granada (Spain). Methods. We have targeted a number of astronomical sources. Starting from beam-maps on primary and secondary calibrators we have then gone to extended sources and faint objects. Both internal (electronic) and on-the-sky calibrations are applied. The general methods are described in the present paper. Results. NIKA2 has been successfully deployed and commissioned, performing in-line with expectations. In particular, NIKA2 exhibits full width at half maximum angular resolutions of around 11 and 17.5 arcsec at respectively 260 and 150 GHz. The noise equivalent flux densities are, at these two respective frequencies, 33±2 and 8±1 mJy s1/2. A first successful science verification run was achieved in April 2017. The instrument is currently offered to the astronomy community and will remain available for at least the following ten years.
The detailed investigation of the broadband flux variability in the blazar 3C 273 allowed us to probe the location and size of emission regions and their physical conditions. We conducted correlation ...studies of the flaring activity in 3C 273, which was observed for the period between 2008 and 2012. The observed broadband variations were investigated using the structure function and the discrete correlation function methods. Starting from the commonly used power spectral density (PSD) analysis at X-ray frequencies, we extended our investigation to characterise the nature of variability at radio, optical, and γ-ray frequencies. The PSD analysis showed that the optical and infrared light-curve slopes are consistent with the slope of white-noise processes, while the PSD slopes at radio, X-ray, and γ-ray energies are consistent with red-noise processes. We found that the estimated fractional variability amplitudes strongly depend on the observed frequency. The flux variations at γ-ray and mm-radio bands are found to be significantly correlated. Using the estimated time lag of (110 ± 27) days between γ-ray and radio light-curves, where γ-ray variations lead the radio bands, we constrained the location of the γ-ray emission region at a de-projected distance of 1.2 ± 0.9 pc from the jet apex. Flux variations at X-ray bands were found to have a significant correlation with variations at both radio and γ-ray energies. The correlation between X-ray and γ-ray light curves indicates two possible time lags, which suggests that two components are responsible for the X-ray emission. A negative time lag of −(50 ± 20) days, where the X-rays are leading the emission, suggests that X-rays are emitted closer to the jet apex from a compact region (0.02–0.05 pc in size), most likely from the corona at a distance of (0.5 ± 0.4) pc from the jet apex. A positive time lag of (110 ± 20) days (γ-rays are leading the emission) suggests a jet-base origin of the other X-ray component at ~4 to 5 pc from the jet apex. The flux variations at radio frequencies were found to be well correlated with each other such that the variations at higher frequencies are leading the lower frequencies, which is expected from the standard shock-in-jet model.
Islet Amyloid Polypeptide (IAPP or amylin) is a peptide hormone produced and stored in the β‐islet cells of the pancreas along with insulin. IAPP readily forms amyloid fibrils in vitro, and the ...deposition of fibrillar IAPP has been correlated with the pathology of type II diabetes. The mechanism of the conversion that IAPP undergoes from soluble to fibrillar forms has been unclear. By chaperoning IAPP through fusion to maltose binding protein, we find that IAPP can adopt a α‐helical structure at residues 8–18 and 22–27 and that molecules of IAPP dimerize. Mutational analysis suggests that this dimerization is on the pathway to fibrillation. The structure suggests how IAPP may heterodimerize with insulin, which we confirmed by protein crosslinking. Taken together, these experiments suggest the helical dimerization of IAPP accelerates fibril formation and that insulin impedes fibrillation by blocking the IAPP dimerization interface.
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