Conformation of [1-13C,15N]Acetyl-L-carnitine. Rotational-Echo, Double-Resonance Nuclear Magnetic Resonance Spectroscopy

Anderson, Robert C.; Gullion, Terry; Joers, James M.; Shapiro, Michael J.; Villhauer, Edwin B.; Weber, Hans Peter

doi:10.1021/ja00147a016

Cited by 50 publications

(90 citation statements)

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“…13 C{ 15 N} rotational-echo double-resonance (REDOR) experiments (21)(22)(23) were performed on a Chemagnetics Infinity 300 spectrometer operating at a magnetic field of 7.05 T, equipped with a 4-mm triple-resonance magic-angle sample spinning probe, with parameters as follows: magic-angle sample spinning frequency 6 kHz, 13 C radio frequency field amplitude of 50 kHz, 15 N radio frequency amplitude of 40 kHz, 1 H decoupling at an amplitude of 110 kHz. The pulse sequence of Anderson et al (24) was used. Between 512 and 2,048 transients per data point were taken on a water-washed and N 2 -dried Ϸ30 mg of fibril sample prepared in 200 mM sodium phosphate, pH 7.2, at 37°C.…”

Section: Methodsmentioning

confidence: 99%

Exploring amyloid formation by a de novo design

Kammerer

Kostrewa²,

Zurdo³

et al. 2004

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

164

194

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Protein deposition as amyloid fibrils underlies many debilitating human disorders. The complexity and size of disease-related polypeptides, however, often hinders a detailed rational approach to study effects that contribute to the process of amyloid formation. We report here a simplified peptide sequence successfully designed de novo to fold into a coiled-coil conformation under ambient conditions but to transform into amyloid fibrils at elevated temperatures. We have determined the crystal structure of the coiled-coil form and propose a detailed molecular model for the peptide in its fibrillar state. The relative stabilities of the two structural forms and the kinetics of their interconversion were found to be highly sensitive to small sequence changes. The results reveal the importance of specific packing interactions on the kinetics of amyloid formation and show the potential of this exceptionally favorable system for probing details of the molecular origins of amyloid disease.

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Section: Methodsmentioning

confidence: 99%

Exploring amyloid formation by a de novo design

Kammerer

Kostrewa²,

Zurdo³

et al. 2004

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

164

194

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“…20,52,[63][64][65] XY-8 phase cycling was applied to the 13 C and 15 N pulses during the dephasing period, TPPM 1 H decoupling of ~95 kHz was applied during the dephasing and detection periods, the recycle delay was 1 s, and the MAS frequency was 8000 ± 2 Hz. REDOR experiments were calibrated using a lyophilized "I4" peptide with sequence AcAE-AAAKEAAAKEAAAKA-NH 2 and a 13 CO label at Ala-9 and a 15 N label at Ala-13.…”

Section: Rotational-echo Double-resonance (Redor) Experiments and Simmentioning

confidence: 99%

Solid-State NMR Spectroscopy of Human Immunodeficiency Virus Fusion Peptides Associated with Host-Cell-Like Membranes: 2D Correlation Spectra and Distance Measurements Support a Fully Extended Conformation and Models for Specific Antiparallel Strand Registries

2008

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The human immunodeficiency virus (HIV) is "enveloped" by a membrane, and infection of a host cell begins with fusion between viral and target cell membranes. Fusion is catalyzed by the HIV gp41 protein which contains a functionally critical ~20-residue apolar "fusion peptide" (HFP) that associates with target cell membranes. In this study, chemically synthesized HFPs were associated with host-cell-like membranes and had "scatter-uniform" labeling (SUL), that is, only one residue of each amino acid type was U-13 C, 15 N labeled. For the first sixteen HFP residues, an unambiguous 13 C chemical shift assignment was derived from 2D 13 C/ 13 C correlation spectra with short mixing times, and the shifts were consistent with continuous β-strand conformation. 13 C-13 C contacts between residues on adjacent strands were derived from correlation spectra with long mixing times and suggested close proximity of the following residues: Ala-6/Gly-10, Ala-6/Phe-11, and Ile-4/Gly-13. Specific antiparallel β-strand registries were further tested using a set of HFPs that were 13 CO-labeled at Ala-14 and 15 N-labeled at either Val-2, Gly-3, Ile-4, or Gly-5. The solid-state NMR data were fit with 50-60% population of antiparallel HFP with either Ala-14/Gly-3 or Ala-14/Ile-4 registries and 40-50% population of structures not specified by the NMR experiments. The first two registries correlated with intermolecular hydrogen bonding of 15-16 apolar N-terminal residues and this hydrogen-bonding pattern would be consistent with a predominant location of these residues in the hydrophobic membrane interior. To our knowledge, these results provide the first residue-specific structural models for membrane-associated HFP in its β-strand conformation.Correspondence to: David P. Weliky, weliky@chemistry.msu.edu. Supporting Information Available: 2D PDSD spectra of HFP-F at 10 and 500 ms exchange times; 1D slices for the 2D PDSD spectra of HFP-C and HFP-D; REDOR spectra at 24 and 32 ms dephasing times for HFP-I and HFP-K samples prepared with 25 µM initial HFP concentration; derivation of (ΔS/S 0 ) cor from (ΔS/S 0 ) exp and σ cor from σ exp ; comparison of (ΔS/S 0 ) exp with (ΔS/S 0 ) cor for the HFP-H, I, J, and K samples; model geometries for the HFP-H, HFP-J, and HFP-K samples; and (ΔS/S 0 ) sim for all models for the HFP-H, HFP-I, HFP-J, and HFP-K samples. This material is available free of charge via the Internet at

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“…Measurements of either 13 C-13 C (Lansbury et al, 1995) or 15 N-13 C Gordon et al, 2004;Kammerer et al, 2004;Petkova et al, 2004) intermolecular dipoledipole couplings can be used to identify antiparallel β-sheets in samples with selective isotopic labeling. Measurements of intermolecular couplings between 15 N-labeled backbone amide sites and 13 C-labeled backbone carbonyl sites, using the rotational echo double resonance (REDOR) recoupling technique (Anderson et al, 1995;Gullion and Schaefer, 1989), are particularly powerful because the 4.1 Å internuclear distance for hydrogen bonded backbone amide and carbonyl sites is significantly shorter than other intermolecular distances in β-sheets. REDOR data such as those in Fig.…”

Section: Data Inmentioning

confidence: 99%

“…Measurements of intermolecular dipole-dipole couplings between 15 N labels at backbone amide sites and 13 C labels at backbone carbonyl sites in amyloid fibrils formed by Aβ [16][17][18][19][20][21][22] with either acetyl (circles, downward triangles) or octanoyl (upward triangles, squares) groups at the N-terminus. These measurements use the REDOR solid state NMR technique (Anderson et al, 1995;Gullion and Schaefer, 1989). Fibrils are formed from mixtures of molecules that are 15 N-labeled at A21 and molecules that are 13 C-labeled at L17 (circles, squares) or F20 (triangles), with an approximate 1.6:1.0 ratio of 15 N-labeled to 13 C-labeled molecules.…”

Section: Sample Preparation For Solid State Nmrmentioning

confidence: 99%

Characterization of Amyloid Structures at the Molecular Level by Solid State Nuclear Magnetic Resonance Spectroscopy

Tycko¹

2006

Methods in Enzymology

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Solid state nuclear magnetic resonance (NMR) spectroscopy is particularly useful in structural studies of amyloid fibrils because solid state NMR techniques have unique capabilities as site-specific, molecular-level structural probes of noncrystalline materials. These techniques provide experimental data that strongly constrain the secondary, tertiary, and quaternary structures of amyloid fibrils, permitting the development of experimentally-based structural models. Examples of techniques that are applicable to amyloid samples prepared with isotopic labeling of specific sites and to samples prepared with uniform isotopic labeling of selected residues are presented, illustrating the utility of the various techniques and labeling schemes. Information regarding the preparation of amyloid samples for solid state NMR measurements is also included.

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Conformation of [1-13C,15N]Acetyl-L-carnitine. Rotational-Echo, Double-Resonance Nuclear Magnetic Resonance Spectroscopy

Cited by 50 publications

References 0 publications

Exploring amyloid formation by a de novo design

Exploring amyloid formation by a de novo design

Solid-State NMR Spectroscopy of Human Immunodeficiency Virus Fusion Peptides Associated with Host-Cell-Like Membranes: 2D Correlation Spectra and Distance Measurements Support a Fully Extended Conformation and Models for Specific Antiparallel Strand Registries

Characterization of Amyloid Structures at the Molecular Level by Solid State Nuclear Magnetic Resonance Spectroscopy

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