X-Ray Diffraction Studies on Amyloid Filaments

Eanes, E. D.; Glenner, G. G.

doi:10.1177/16.11.673

Cited by 721 publications

(522 citation statements)

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“…Both samples were prepared from AMY05 and were hydrated in these measurements. 13 C chemical shifts at the positions of maximum crosspeak intensity are similar in the two spectra, with the exception of chemical shifts for Ala5, for which the C β chemical shifts differ by 8 ppm, and Ile26, for which the C δ chemical shifts differ by 2 ppm. However, linewidths in the spectrum of polymorphic fibrils are greater than 2.5 ppm (full width at half maximum, determined from crosspeaks in the 2D spectra), while linewidths in the spectrum of morphologically homogeneous fibrils range from 1.0 ppm to 2.0 ppm and are typically less than 1.5 ppm.…”

Section: Characterization Of Secondary Structure By 13 C Nmrmentioning

confidence: 81%

Section: Solid State Nmrmentioning

confidence: 99%

“…In 2D radiofrequency-assisted diffusion (RAD) measurements, exchange periods ranging from 500 ms to 1000 ms were used, during which an rf field amplitude equal to the MAS frequency was applied to protons (61,62). With long exchange periods, strong crosspeaks in 2D RAD spectra connect all 13 C NMR lines arising from a single labeled residue or from pairs of labeled residues that occur sequentially (i.e., consecutively in the amylin sequence). Weaker inter-residue crosspeaks arising from contacts between nonsequential residue pairs provide constraints on peptide conformation.…”

Section: Solid State Nmrmentioning

confidence: 99%

“…In the second step, two copies of a pentamer (i.e., a final structure from the first step) were manually rotated and displaced such that the β-sheet segments comprised of residues 28-37 were antiparallel and about 20 Å apart, producing a two-fold symmetric structure consistent with experimental STEM data and solid state 13 C NMR spectra (see below). In addition to the restraints within each pentamer, intermolecular restraints between C α atoms of Leu27-Val32 pairs were added to attract the two pentamers to one another.…”

Section: Construction Of Molecular Modelsmentioning

confidence: 99%

“…In 2D finite-pulse radiofrequency-driven recoupling (fpRFDR) measurements, the fpRFDR sequence (16,60) was applied during exchange periods of 1.5 or 2.2 ms between the t 1 and t 2 dimensions, with 13 C π pulse lengths set to 30% of the MAS rotation period. Strong crosspeaks in the 2D fpRFDR spectra connect 13 C NMR lines of directly-bonded carbon sites. In 2D radiofrequency-assisted diffusion (RAD) measurements, exchange periods ranging from 500 ms to 1000 ms were used, during which an rf field amplitude equal to the MAS frequency was applied to protons (61,62).…”

Section: Solid State Nmrmentioning

confidence: 99%

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Peptide Conformation and Supramolecular Organization in Amylin Fibrils: Constraints from Solid-State NMR

et al. 2007

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The 37-residue amylin peptide, also known as islet amyloid polypeptide, forms fibrils that are the main peptide or protein component of amyloid that develops in the pancreas of type 2 diabetes patients. Amylin also readily forms amyloid fibrils in vitro that are highly polymorphic under typical experimental conditions. We describe a protocol for the preparation of synthetic amylin fibrils that exhibit a single predominant morphology, which we call a striated ribbon, in electron microscope and atomic force microscope images. Solid state nuclear magnetic resonance (NMR) measurements on a series of isotopically labeled samples indicate a single molecular structure within the striated ribbons. We use scanning transmission electron microscopy and several types of one-dimensional and two-dimensional solid state NMR techniques to obtain constraints on the peptide conformation and supramolecular structure in these amylin fibrils, and derive molecular structural models that are consistent with the experimental data. The basic structural unit in amylin striated ribbons, which we call the protofilament, contains four-layers of parallel β-sheets, formed by two symmetric layers of amylin molecules. The molecular structure of amylin protofilaments in striated ribbons closely resembles the protofilament in amyloid fibrils with similar morphology formed by the 40-residue β-amyloid peptide that is associated with Alzheimer's disease. Keywordsislet amyloid polypeptide; type 2 diabetes; amyloid fibril structure; electron microscopy; atomic force microscopy Amylin, also called islet amyloid polypeptide or IAPP, is a 37-residue peptide that is cosecreted with insulin by the β-cells of pancreas. The normal biological effects of amylin secretion include inhibition of glucagon secretion and reduction of the rate of gastric emptying, effects that contribute to the maintenance of postprandial glucose homeostasis (1). Amylin is the major peptide or protein component of the islet amyloid found in the pancreas of approximately 90% of type 2 (or non-insulin-dependent) diabetes patients (2,3). Fibrillar amylin has been shown * Corresponding author: Dr. Robert Tycko, National Institutes of Health, Building 5, Room 112, Bethesda, MD 20892-0520. phone 301-402-8272. fax 301-496-0825. e-mail robertty@mail.nih.gov.Supporting Information Available HPLC chromatograms from the purification of both reduced and oxidized amylin by reverse-phase chromatography ( Figure S1); FPLC chromatograms from the purification of both human and rodent amylin by size-exclusion chromatography ( Figure S2); 1D 13 C spectra of amylin fibrils with various isotopic labeling patterns, in lyophilized and rehydrated conditions ( Figures S3-S5); Table of 13 C NMR chemical shifts in amylin fibrils, TALOS predictions of backbone torsion angles based on these shifts, and torsion angles determined from 15 N fpRFDR-CT measurements (Table S1). This material is freely available on the World Wide Web at http://www.acs.org. to be toxic to cultured β-cells (4), and has been propo...

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Section: Characterization Of Secondary Structure By 13 C Nmrmentioning

confidence: 81%