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  • Amyloid fibrils are insoluble high molecular

    2024-04-03

    Amyloid fibrils are insoluble, high-molecular-weight, non-crystalline aggregates and traditional experimental techniques for structure determination such as X-ray and solution NMR cannot resolve their high resolution structures [64], [68]. However, amyloid fibrils exhibit various advantageous features for solid state NMR. 15N and 13C atoms in structured regions have relatively small line widths (0.5–2.5 ppm) and also favorable nuclear spin relaxation properties. Furthermore, fibril samples typically have high protein or peptide concentrations (25–100%, depending on hydration level), which permits use of small sample volumes (5–50 μL) necessary for fast magic angle spinning (MAS). Production of isotopically labeled polypeptides is quite feasible both by solid phase peptide synthesis and bacterial expression methods [70], [148]. Moreover, protocols have been published for generating homogenous fibril samples displaying excellent solid state NMR spectra with sharp resonance lines [149], [150]. Various NMR techniques are applied to provide a broad range of constraints for characterizing the secondary, tertiary and quaternary structure of fibrils. Secondary structure determination in fibrils refers to β-strand and non-β-strand segment identification as fibrils containing α-helical structures have not been reported. Moreover, these constraints can resolve the organization of β-sheets into parallel or antiparallel and the register of interstrand hydrogen bonds and provide data on the relative orientation of β-sheets and interresidue contacts of amino 5003 sale side chains. Generally, ssNMR experiments for acquiring distance constraints utilize measurements of dipole-dipole interactions between nuclei. For instance, proton assisted recoupling (PAR) [151], [152], dipolar assisted rotational resonance (DARR) [153] and frequency selective rotational echo double resonance (FS-REDOR) [154], [155], [156] are some frequently used pulse sequences to obtain 13C-13C or 13C-15N long range constraints. Although ssNMR provides a large number of constraints, complementary techniques can supply data on the overall topology of fibrils and significantly simplify determination of the structure. Scanning TEM measurements can determine the mass-per-length (MPL) of fibrils, which is an extremely important constraint on 5003 sale the quaternary structure [157].
    Conclusions We have reviewed here current knowledge on the structures of various Aβ species starting from monomers to mature fibrils. Based on these experimental results we have discussed the possible reasons for the striking differences in both aggregation rates and toxicity between Aβ1-40 and Aβ1-42. The task of elucidating the aggregation mechanism is complicated by the observed polymorphism for both oligomers and fibrils. Most likely every fibril polymorph has a disparate aggregation pathway. Nevertheless, familiar structural properties can be identified between different Aβ1-40 fibril polymorphs and perhaps a universal mechanism of aggregation can be derived. In contrast, Aβ1-40 and Aβ1-42 probably cannot be described by the same aggregation pathway since they show very different fibril structures. Both Aβ alloforms may exhibit a different aggregation pathway from a structural perspective, however they involve the same aggregation kinetic stages. In this review primary nucleation mechanisms have been considered that provide an explanation on the 100-fold faster nucleation rate of Aβ1-42. The higher secondary nucleation rate of Aβ1-42 can be construed as a consequence of extra surface exposed hydrophobic patches of Aβ1-42 fibrils. Nevertheless, further research is still required to determine the exact aggregation mechanism for each alloform, especially important are structural studies of on-pathway intermediate aggregates. Currently, almost all of the characterized oligomer structures involve a β-strand-turn-β-strand motif similar to that of the Aβ1-40 fibril. However, these intermediate states are unlikely to be on-pathway to Aβ1-42 fibril formation.