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           dipolar recoupling under very fast magic angle spinning in solid-state nuclear magnetic resonance: Applications to distance measurements, spectral assignments, and high-throughput secondary-structure determination

Ishii, Yoshitaka

doi:10.1063/1.1359445

Cited by 272 publications

(280 citation statements)

References 70 publications

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“…Fig. 2 shows two-dimensional (2D) 13 C-13 C NMR spectra of the five SFg2 samples, obtained with 2.4 ms finite-pulse radio-frequency-driven recoupling (fpRFDR) mixing periods (35,36) so that strong crosspeaks connect chemical shifts of directly bonded, 13 C-labeled sites. Except for F19 and A21, all residues show a single set of chemical shifts, consistent among samples A-E, indicating that all D23N-Aβ 1-40 molecules have similar conformations and structural environments and that sample preparation is reproducible.…”

Section: Resultsmentioning

confidence: 99%

Antiparallel β-sheet architecture in Iowa-mutant β-amyloid fibrils

Wang

Yau

Luo

et al. 2012

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

329

533

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Wild-type, full-length (40-and 42-residue) amyloid β-peptide (Aβ) fibrils have been shown by a variety of magnetic resonance techniques to contain cross-β structures in which the β-sheets have an in-register parallel supramolecular organization. In contrast, recent studies of fibrils formed in vitro by the Asp23-to-Asn mutant of 40-residue Aβ (D23N-Aβ 1-40 ), which is associated with early onset neurodegeneration, indicate that D23N-Aβ 1-40 fibrils can contain either parallel or antiparallel β-sheets. We report a protocol for producing structurally pure antiparallel D23N-Aβ 1-40 fibril samples and a series of solid state nuclear magnetic resonance and electron microscopy measurements that lead to a specific model for the antiparallel D23N-Aβ 1-40 fibril structure. This model reveals how both parallel and antiparallel cross-β structures can be constructed from similar peptide monomer conformations and stabilized by similar sets of interactions, primarily hydrophobic in nature. We find that antiparallel D23N-Aβ 1-40 fibrils are thermodynamically metastable with respect to conversion to parallel structures, propagate less efficiently than parallel fibrils in seeded fibril growth, and therefore must nucleate more efficiently than parallel fibrils in order to be observable. Experiments in neuronal cell cultures indicate that both antiparallel and parallel D23N-Aβ 1-40 fibrils are cytotoxic. Thus, our antiparallel D23N-Aβ 1-40 fibril model represents a specific "toxic intermediate" in the aggregation process of a disease-associated Aβ mutant.Alzheimer's disease | amyloid structure | solid state NMR A lzheimer's disease (AD) is thought to be a consequence of aggregation of the amyloid β-peptide (Aβ) into amyloid fibrils or related assemblies in brain tissue. Numerous structural studies of Aβ fibrils and oligomers have been reported (1-17), motivated by the dual goals of contributing to preventive and therapeutic approaches to AD and of elucidating the biophysical basis for amyloid formation. A defining structural characteristic of an amyloid fibril is the presence of a cross-β motif; i.e., a ribbon-like β-sheet running the length of the fibril, with β-strands approximately perpendicular to and interstrand hydrogen bonds approximately parallel to the long fibril axis. Studies of the 40-residue and 42-residue forms of Aβ (Aβ 1-40 and Aβ 1-42 ) have shown that these peptides can form multiple distinct fibril structures (11,18,19), but that the cross-β motifs within wild-type (WT) Aβ fibrils are invariably comprised of in-register parallel β-sheets (1, 5, 6, 8-10, 13, 14). Parallel β-sheets have also been found in Aβ 1-40 oligomers (4) and in amyloid fibrils formed by amylin (20, 21), α-synuclein (22), β 2 -microglobulin (23, 24), prion proteins of yeast (25-27) and mammalian PrP (28, 29); in contrast, antiparallel β-sheets have been found in fibrils formed by Aβ fragments with 15 or fewer residues (30-32) and in amyloid-like crystals of certain Aβ fragments (33). These observations suggest that in-register parallel β-shee...

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Section: Resultsmentioning

confidence: 99%

Antiparallel β-sheet architecture in Iowa-mutant β-amyloid fibrils

Wang

Yau

Luo

et al. 2012

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

329

533

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“…The sample was measured by 1 H-13 C cross-polarization in conjunction with finite pulse radio frequency-driven recoupling (35) or radio frequency-assisted diffusion (36,37) pulse sequences to detect solidstate NMR signals and correlate 13 C resonance frequencies of bonded or spatially close carbons (e.g. in the same amino acid residue).…”

Section: Methodsmentioning

confidence: 99%

Repeat Domains of Melanosome Matrix Protein Pmel17 Orthologs Form Amyloid Fibrils at the Acidic Melanosomal pH

McGlinchey¹,

Shewmaker²,

et al. 2011

Journal of Biological Chemistry

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Most amyloids are pathological, but fragments of Pmel17 form a functional amyloid in vertebrate melanosomes essential for melanin synthesis and deposition. We previously reported that only at the mildly acidic pH (4 -5.5) typical of melanosomes, the repeat domain (RPT) of human Pmel17 can form amyloid in vitro. Combined with the known presence of RPT in the melanosome filaments and the requirement of this domain for filament formation, we proposed that RPT may be the core of the amyloid formed in vivo. Although most of Pmel17 is highly conserved across a broad range of vertebrates, the RPT domains vary dramatically, with no apparent homology in some cases. Here, we report that the RPT domains of mouse and zebrafish, as well as a small splice variant of human Pmel17, all form amyloid specifically at mildly acid pH (pH ϳ5.0). Protease digestion, mass per unit length measurements, and solid-state NMR experiments suggest that amyloid of the mouse RPT has an in-register parallel ␤-sheet architecture with two RPT molecules per layer, similar to amyloid of the A␤ peptide. Although there is no sequence conservation between human and zebrafish RPT, amyloid formation at acid pH is conserved.Amyloid is a filamentous ␤-sheet-rich protein polymer usually associated with Alzheimer disease, Parkinson disease, type II diabetes, and other pathological conditions, but several functional amyloids have been identified. The external curli fibers of Escherichia coli are involved in biofilm formation and adhesion to host cells (1); the hydrophobin amyloids protect fungal cell surfaces (2), and the [Het-s] prion of Podospora anserina appears to help the host by facilitating heterokaryon incompatibility (3).Pmel17 is a protein important for melanin synthesis and deposition that forms intralumenal filaments in melanosomes, the endosome/lysosome-like organelle in which melanin synthesis takes place (reviewed in Refs. 4, 5). Pmel17 is made as a transmembrane protein, processed by a series of proteolytic steps and glycosylations and assembled into a filamentous form in stage II melanosomes. There, Pmel17 filaments facilitate melanization, perhaps acting as a template on which melanin is deposited or by adsorbing toxic by-products of the melanin biosynthetic process (reviewed in Ref. 6).The gene for Pmel17 was first identified by a single gene mouse mutant called Silver, in which the normal black coat color was lost as animals grew and aged (7). The mouse Pmel17 cDNA, mapping at the Silver locus, defined the protein with its single transmembrane region and repeat domain (8). Comparisons of orthologs from a number of vertebrate species have revealed the domain structure shown in supplemental Fig. 1 (6, 9). Human Pmel17 consists of (N-to C-terminal) a signal peptide (residues 1-23 in the human protein), an N-terminal domain, the polycystic kidney disease-like domain (PKD), 2 the repeat domain (RPT), the Kringle-like domain (KLD or KRG), the transmembrane domain, and the cytoplasmic C-terminal domain. Most parts of Pmel17 are well aligned acros...

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“…[1][2][3][4][5][6][7][8][9][10][11][12][13][14][15][16] Particularly, use of protein micro-/nano-crystals [17] has significantly improved resolution in high-resolution SSNMR of dilute spins such as 13 C and 15 N, permitting signal assignment and structural determination of various uniformly 13 C and/or 15 N-labeled proteins by SSNMR. [18][19][20][21][22][23][24][25] However, restricted sensitivity in 13 C and 15 N SSNMR has been still one of the major limiting factors in SSNMR analysis of proteins.…”

Section: Introductionmentioning

confidence: 99%

Sensitivity enhancement in 13C solid-state NMR of protein microcrystals by use of paramagnetic metal ions for optimizing 1H T1 relaxation

Wickramasinghe

Kotecha

Samoson

et al. 2007

Journal of Magnetic Resonance

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117

110

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We discuss a simple approach to enhance sensitivity for 13 C high-resolution solid-state NMR for proteins in microcrystals by reducing 1 H T 1 relaxation times with paramagnetic relaxation reagents. It was shown that 1 H T 1 values can be reduced from 0.4-0.8 s to 60-70 ms for ubiquitin and lysozyme in D 2 O in the presence of 10 mM Cu(II)Na 2 EDTA without substantial degradation of the resolution in 13 C CPMAS spectra. Faster signal accumulation using the shorter 1 H T 1 attained by paramagnetic doping provided sensitivity enhancements of 1.4-2.9 for these proteins, reducing the experimental time for a given signal-to-noise ratio by a factor of 2.0-8.4. This approach presented here is likely to be applicable to various other proteins in order to enhance sensitivity in 13 C high-resolution solidstate NMR spectroscopy.

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C 13 – C 13 dipolar recoupling under very fast magic angle spinning in solid-state nuclear magnetic resonance: Applications to distance measurements, spectral assignments, and high-throughput secondary-structure determination

Cited by 272 publications

References 70 publications

Antiparallel β-sheet architecture in Iowa-mutant β-amyloid fibrils

Antiparallel β-sheet architecture in Iowa-mutant β-amyloid fibrils

Repeat Domains of Melanosome Matrix Protein Pmel17 Orthologs Form Amyloid Fibrils at the Acidic Melanosomal pH

Sensitivity enhancement in 13C solid-state NMR of protein microcrystals by use of paramagnetic metal ions for optimizing 1H T1 relaxation

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