Carbon-detected deuterium solid-state NMR rotating frame relaxation measurements for protein methyl groups under magic angle spinning.

Deuterium rotating frame solid-state NMR relaxation measurements (2H R1ρ) are important tools in quantitative studies of molecular dynamics. We demonstrate how 2H to 13C cross-polarization (CP) approaches under 10-40 kHz magic angle spinning rates can be combined with the 2H R1ρ blocks to allow for extension of deuterium rotating frame relaxation studies to methyl groups in biomolecules. This extension permits detection on the 13C nuclei and, hence, for the achievement of site-specific resolution. The measurements are demonstrated using a nine-residue low complexity peptide with the sequence GGKGMGFGL, in which a single selective -13CD3 label is placed at the methionine residue. Carbon-detected measurements are compared with the deuterium direct-detection results, which allows for fine-tuning of experimental approaches. In particular, we show how the adiabatic respiration CP scheme and the double adiabatic sweep on the 2H and 13C channels can be combined with the 2H R1ρ relaxation rates measurement. Off-resonance 2H R1ρ measurements are investigated in addition to the on-resonance condition, as they extent the range of effective spin-locking field.

Rigidifying of the internal dynamics of amyloid-beta fibrils generated in the presence of synaptic plasma vesicles.

We investigated the changes in internal flexibility of amyloid-ß1-40 (Aß) fibrils grown in the presence of rat synaptic plasma vesicles. The fibrils are produced using a modified seeded growth protocol, in which the Aß concentration is progressively increased at the expense of the decreased lipid to protein ratio. The morphologies of each generation are carefully assessed at several fibrils' growth time points using transmission electron microscopy. The side-chain dynamics in the fibrils is investigated using deuterium solid-state NMR measurements, with techniques spanning line shapes analysis and several NMR relaxation rates measurements. The dynamics is probed in the site-specific fashion in the hydrophobic C-terminal domain and the disordered N-terminal domain. An overall strong rigidifying effect is observed in comparison with the wild-type fibrils generated in the absence of the membranes. In particular, the overall large-scale fluctuations of the N-terminal domain are significantly reduced, and the activation energies of rotameric inter-conversion in methyl-bearing side-chains of the core (L17, L34, M35, V36), as well as the ring-flipping motions of F19 are increased, indicating a restricted core environment. Membrane-induced flexibility changes in Aß aggregates can be important for the re-alignment of protein aggregates within the membrane, which in turn would act as a disruption pathway of the bilayers' integrity.

Persistence of Methionine Side Chain Mobility at Low Temperatures in a Nine-Residue Low Complexity Peptide, as Probed by 2 H Solid-State NMR.

Methionine side chains are flexible entities which play important roles in defining hydrophobic interfaces. We utilize deuterium static solid-state NMR to assess rotameric inter-conversions and other dynamic modes of the methionine in the context of a nine-residue random-coil peptide (RC9) with the low-complexity sequence GGKGMGFGL. The measurements in the temperature range of 313 to 161 K demonstrate that the rotameric interconversions in the hydrated solid powder state persist to temperatures below 200 K. Removal of solvation significantly reduces the rate of the rotameric motions. We employed 2 H NMR line shape analysis, longitudinal and rotation frame relaxation, and chemical exchange saturation transfer methods and found that the combination of multiple techniques creates a significantly more refined model in comparison with a single technique. Further, we compare the most essential features of the dynamics in RC9 to two different methionine-containing systems, characterized previously. Namely, the M35 of hydrated amyloid-ß1-40 in the three-fold symmetric polymorph as well as Fluorenylmethyloxycarbonyl (FMOC)-methionine amino acid with the bulky hydrophobic group. The comparison suggests that the driving force for the enhanced methionine side chain mobility in RC9 is the thermodynamic factor stemming from distributions of rotameric populations, rather than the increase in the rate constant.

Surgical Approaches to Petrous Apex Cholesterol Granulomas: A Systematic Review and Network Meta-analysis.

OBJECTIVE: To compare the outcomes of different surgical approaches to petrous apex cholesterol granulomas (PACG). DATA SOURCES: PubMed, Embase, Google Scholar, Cochrane, and Web of Science. REVIEW METHODS: Following the Preferred Reporting Items for Systematic Reviews and Meta-analyses-Network Meta-analyses guidelines, databases were searched from inception to November 31, 2022. Studies comparing two or more approaches were included. Reviews and population studies were excluded. The main outcome measures were the resolution of symptoms, serviceable hearing, complication, and revision rates. RESULTS: The search yielded 2132 studies. After applying inclusion and exclusion criteria, 15 studies remained, consisting of 214 patients treated with lateral approaches (n = 182) or anterior endonasal approaches (n = 32). The efficacy of lateral and anterior endonasal approaches in achieving symptom resolution was comparable (73% vs. 68%, p = 0.5). Both exhibited similar rates of complications (33% vs. 37%, p = 0.3), albeit with distinct profiles. Lateral approaches were associated with higher rates of facial palsy and sensorineural hearing loss (44% vs. 18%, p = 0.03). Anterior endonasal approaches demonstrated higher rates of epistaxis and cerebrospinal fluid leak (15% vs. 1%, p = 0.001). Anterior endonasal approaches exhibited lower revision rates (OR: 0.35, 95% CI: 0.14-0.88). The placement of a stent in both approaches was associated with higher symptom resolution (OR: 5.12, 95% CI: 1.05-9.97) and lower revision rates (OR: 0.71, 95% CI: 0.33-0.92). CONCLUSIONS: Anterior endonasal approaches yield lower revision rates compared to lateral approaches for PACG. Both approaches demonstrate similar effectiveness in symptom resolution and comparable rates of complications, with distinct profiles. Facial nerve and hearing status are important factors that should be addressed when selecting the approach. Stenting is beneficial. LEVEL OF EVIDENCE: N/A Laryngoscope, 2023.

Modulation of aggregation and structural polymorphisms of ß-amyloid fibrils in cellular environments by pyroglutamate-3 variant cross-seeding.

Amyloidogenic deposition of ß-amyloid (Aß) peptides in human brain involves not only the wild-type Aß (wt-Aß) sequences, but also posttranslationally modified Aß (PTM-Aß) variants. Recent studies hypothesizes that the PTM-Aß variants may trigger the deposition of wt-Aß, which underlies the pathology of Sporadic Alzheimer's disease. Among PTM-Aß variants, the pyroglutamate-3-Aß (pyroE3-Aß) has attracted much attention because of their significant abundances and broad distributions in senile plaques and dispersible and soluble oligomers. pyroE3-specific antibodies are being tested as potential anti-Aß drugs in clinical trials. However, evidence that support the triggering effect of pyroE3-Aß on wt-Aß in cells remain lacking, which diminishes its pathological relevance. We show here that cross-seeding with pyroE3-Aß40 leads to accelerated extracellular and intracellular aggregation of wt-Aß40 in different neuronal cells. Cytotoxicity levels are elevated through the cross-seeded aggregation, comparing with the self-seeded aggregation of wt-Aß40 or the static presence of pyroE3-Aß40 seeds. For the extracellular deposition in mouse neuroblastoma Neuro2a (N2a) cells, the cytotoxicity elevation correlates positively with the seeding efficiency. Besides aggregation rates, cross-seeding with pyroE3-Aß40 also modulates the molecular level structural polymorphisms of the resultant wt-Aß40 fibrils. Using solid-state nuclear magnetic resonance (ssNMR) spectroscopy, we identified key structural differences between the parent pyroE3/ΔE3 and wt-Aß40 fibrils within their fibrillar cores. Structural propagation from seeds to daughter fibrils is demonstrated to be more pronounced in the extracellular seeding in N2a cells by comparing the ssNMR spectra from different seeded wt-Aß40 fibrils, but less significant in the intracellular seeding process in human neuroblastoma SH-SY5Y cells.

Deuteron off-resonance rotating frame relaxation for the characterization of slow motions in rotating and static solid-state proteins.

We demonstrate the feasibility of deuterium solid-state NMR off-resonance rotating frame relaxation measurements for studies of slow motions in biomolecular solids. The pulse sequence, which includes adiabatic pulses for magnetization alignment, is illustrated for static and magic-angle spinning conditions away from rotary resonances. We apply the measurements for three systems with selective deuterium labels at methyl groups: a) a model compound, Fluorenylmethyloxycarbonyl methionine-D3 amino acid, for which the principles of the measurements and corresponding motional modeling based on rotameric interconversions are demonstrated; b) amyloid-ß1-40 fibrils labeled at a single alanine methyl group located in the disordered N-terminal domain. This system has been extensively studied in prior work and here serves as a test of the method for complex biological systems. The essential features of the dynamics consist of large-scale rearrangements of the disordered N-terminal domain and the conformational exchange between the free and bound forms of the domain, the latter one due to transient interactions with the structured core of the fibrils. and c) a 15-residue helical peptide which belongs to the predicted α-helical domain near the N-terminus of apolipoprotein B. The peptide is solvated with triolein and incorporates a selectively labeled leucine methyl groups. The method permits model refinement, indicating rotameric interconversions with a distribution of rate constants.

Adult tonsillectomy-increased pain scores are correlated with risk of bleeding: a retrospective cohort study.

PURPOSE: Tonsillectomy is among the most common surgical procedures performed worldwide, and post-tonsillectomy bleeding is a serious complication. This study aims to investigate the role of post-operative pain as a risk factor for bleeding in adults. METHODS: A retrospective cohort study of adults who underwent tonsillectomy in a tertiary referral center between 2015-2021. Medical records were reviewed for demographics, diagnoses, surgical technique, treatments, pain scores (measured by visual analogue scale 0-10), readmissions, and bleeding events. The primary outcome was return to the operating room for hemostasis, and secondary outcomes were bleeding events and consumption of additional analgesic doses. RESULTS: Of the 274 patients, 137 (50%) were males, the mean age was 30.3 ± 12 years (range 18-82), and 33 (12%) were smokers. Indications for tonsillectomy were recurrent throat infections in 213 (77.7%) patients and obstructive sleep apnea in 61 (22.3%). Surgical technique was cold dissection in 238 (86.9%) patients and electrocautery in 36 (13.1%). Primary post-tonsillectomy bleeding (< 24 h of surgery) occurred in 6 (2%) patients, and secondary bleeding (later than 24 h from tonsillectomy) in 43 (15.7%). A total of 19 (7%) patients necessitated surgical hemostasis. After controlling for technique and other confounders, high pain scores (VAS ≥ 5) on post-operative days 1 and 2 were associated with increased risk of bleeding that necessitated surgical hemostasis (adjusted odds ratio 6.9, 95% confidence interval 1.7-44.5). Other independent risk factors were male sex, age < 30 years, smoking, and recurrent throat infections. CONCLUSIONS: Higher pain scores following tonsillectomy are correlated with bleeding episodes requiring surgical intervention in adults. Further studies may explore the role of different intensive pain regimens in minimizing the risk of bleeding.

Effect of Cross-Seeding of Wild-Type Amyloid-ß1-40 Peptides with Post-translationally Modified Fibrils on Internal Dynamics of the Fibrils Using Deuterium Solid-State NMR.

Post-translationally modified (PTM) amyloid-ß (Aß) species can play an important role in modulating Alzheimer's disease pathology. These relatively less populated modifications can cross-seed the wild-type Aß peptides to produce fibrils that retain many structural and functional features of the original PTM variants. We focus on studies of internal flexibility in the cross-seeded Aß1-40 fibrils originating from seeding with two PTM variants with modifications in the disordered N-terminal domain: ΔE3 truncation and S8-phosphorylation. We employ an array of 2H solid-state NMR techniques, including line shape analysis over a broad temperature range, longitudinal relaxation, and quadrupolar CPMG, to assess the dynamics of the cross-seeded fibrils. The focus is placed on selected side-chain sites in the disordered N-terminal domain (G9 and V12) and hydrophobic core methyl and aromatic groups (L17, L34, M35, V36, and F19). We find that many of the essential features of the dynamics present in the original PTM seeds persist in the cross-seeded fibrils, and several of the characteristic features are even enhanced. This is particularly true for the activation energies of the rotameric motions and large-scale rearrangements of the N-terminal domain. Thus, our results on the dynamics complement prior structural and cell toxicity studies, suggesting that many PTM Aß species can aggressively cross-seed the wild-type peptide in a manner that propagates the PTM's signature.

Nine-residue low-complexity disordered peptide as a model system, an NMR/CD study.

Disordered proteins and protein segments can be crucial for biological function. In this work we present a detailed biophysical characterization of the low-complexity nine-residue peptide with the sequence GGKGMGFGL. Based on proton solution NMR chemical shifts, circular dichroism measurements, as well as the analysis of concentration dependence of NMR linewidth, proton longitudinal relaxation times, hydrogen-deuterium exchange measurements, and 15N rotating frame NMR relaxation measurements, we conclude that the peptide is fully disordered and monomeric in solution. The peptide will serve as a model system for future structural and dynamics studies of biologically relevant disordered peptides in solution and solid states.

Slow methyl axes motions in perdeuterated villin headpiece subdomain probed by cross-correlated NMR relaxation measurements.

Protein methyl groups can participate in multiple motional modes on different time scales. Sub-nanosecond to nano-second time scale motions of methyl axes are particularly challenging to detect for small proteins in solutions. In this work we employ NMR relaxation interference between the methyl H-H/H-C dipole-dipole interactions [Sun&Tugarinov, J. Magn. Reason. 2012] to characterize methyl axes motions as a function of temperature in a small model protein villin headpiece subdomain (HP36), in which all non-exchangeable protons are deuterated with the exception of methyl groups of leucine and valine residues. The data points to the existence of slow motional modes of methyl axes on sub-nanosecond to nanosecond time scales. Further, at high temperatures for which the overall tumbling of the protein is on the order of 2 ns, we observe a coupling between the slow internal motion and the overall molecular tumbling, based on the anomalous order parameters and their temperature-dependent trends. The addition of 28%(w/w) glycerol-d8 increases the viscosity of the solvent and separates the timescales of internal and overall tumbling, thus permitting for another view of the necessity of the coupling assumption for these sites at high temperatures.

Deuteron rotating frame relaxation for the detection of slow motions in rotating solids.

We demonstrate experimental and computational approaches for measuring 2H rotating frame NMR relaxation for solid samples under magic angle spinning (MAS) conditions. The relaxation properties of the deuterium spin-1 system are dominated by the reorientation of the anisotropic quadrupolar tensors, with the effective quadrupolar coupling constant around 55 kHz for methyl groups. The technique is demonstrated using the model compound dimethyl-sulfone at MAS rates of 10 and 60 kHz as well as for an amyloid fibril sample comprising an amyloid-ß (1-40) protein with a selective methyl group labeled in the disordered domain of the fibrils, at an MAS rate of 8 kHz. For both systems, the motional parameters fall well within the ranges determined by other techniques, thus validating its feasibility. Experimental and computational factors such as i) the probe's radio frequency inhomogeneity profiles, ii) rotary resonances at conditions for which the spin-lock field strength matches the half- or full-integer of the MAS rate, iii) the choice of MAS rates and spin-lock field strengths, and iv) simulations that account for the interconversion of multiple coherences for the spin-1 system under MAS and deviations from the analytical Redfield treatment are thoroughly considered.

Comparative Hydrophobic Core Dynamics Between Wild-Type Amyloid-ß Fibrils, Glutamate-3 Truncation, and Serine-8 Phosphorylation.

Post-translational modifications (PTMs) of amyloid-ß (Aß) species are implicated in the modulation of overall toxicities and aggregation propensities. We investigated the internal dynamics in the hydrophobic core of the truncated ΔE3 mutant fibrils of Aß1-40 and compared them with prior and new data for wild-type fibrils as well as with phosphorylated S8 fibrils. Deuteron static solid-state NMR techniques, spanning line-shape analysis, longitudinal relaxation, and chemical exchange saturation transfer methods, were employed to assess the rotameric jumps of several methyl-bearing and aromatic groups in the core of the fibrils. Taken together, the results indicate the rather significant influence of the PTMs on the hydrophobic core dynamics, which propagates far beyond the local site of the chemical modification. The phosphorylated S8 fibrils display an overall rigidifying of the core based on the higher activation barriers of motions than the wild-type fibrils, whereas the ΔE3 fibrils induce a broader variety of changes, some of which are thermodynamic in nature rather than the kinetic ones.

Mental health clinicians' attitudes toward narcissistic personality disorder.

Narcissistic personality disorder (NPD) is an underdiagnosed psychiatric condition in which there is minimal research to support a validated treatment regimen. As a result, it is unclear how mental health practitioners approach NPD patients and the outcomes of clinical management. Given this gap in current mental health practice, this article explores the attitudes of clinicians toward treating NPD patients and the clinician-reported outcomes of managing NPD patients. The study uses a qualitative and quantitative approach to analyze the results of a 16-question survey about clinicians' experiences working with NPD patients. 173 participants were recruited from online psychology interest forums between April 2019 to August 2019. The data were analyzed using the Statistical Package for the Social Scientist (SPSS-26). The qualitative data showed that clinicians find NPD patients to be difficult and challenging. Additionally, clinicians report minimal experience treating NPD and high treatment drop-out rates. The quantitative data were evaluated using Pearson correlation coefficients. These results show that clinicians who view NPD as a diagnosis worth treating report more benefit from their care. These results also showed that participants who report formal didactic training in NPD have better results with their patients. The data also show that clinicians feel group therapy can be helpful in treating NPD, despite reporting that it is often not offered to NPD patients. This is an area that can direct future NPD research to develop a comprehensive approach to NPD management. (PsycInfo Database Record (c) 2021 APA, all rights reserved).

Deuteron Chemical Exchange Saturation Transfer for the Detection of Slow Motions in Rotating Solids.

We utilized the 2H Chemical Exchange Saturation Transfer (CEST) technique under magic angle spinning (MAS) conditions to demonstrate the feasibility of the method for studies of slow motions in the solid state. For the quadrupolar anisotropic interaction, the essence of CEST is to scan the saturation pattern over a range of offsets corresponding to the entire spectral region(s) for all conformational states involved, which translates into a range of -60-+ 60 kHz for methyl groups. Rotary resonances occur when the offsets are at half-and full-integer of the MAS rates. The choice of the optimal MAS rate is governed by the condition to reduce the number of rotary resonances in the CEST profile patterns and retain a sufficiently large quadrupolar interaction active under MAS to maintain sensitivity to motions. As examples, we applied this technique to a well-known model compound dimethyl-sulfone (DMS) as well as amyloid-ß fibrils selectively deuterated at a single methyl group of A2 belonging to the disordered domain. It is demonstrated that the obtained exchange rate between the two rotameric states of DMS at elevated temperatures fell within known ranges and the fitted model parameters for the fibrils agree well with the previously obtained value using static 2H NMR techniques. Additionally, for the fibrils we have observed characteristic broadening of rotary resonances in the presence of conformational exchange, which provides implications for model selection and refinement. This work sets the stage for future potential extensions of the 2H CEST under MAS technique to multiple-labeled samples in small molecules and proteins.

Long-Term Quality of Life after Endoscopic Pituitary Adenoma Surgery with Nasoseptal Flap Reconstruction.

INTRODUCTION: Endoscopic endonasal transsphenoidal surgery (EETS) on the pituitary gland is considered safe and efficacious. The nasoseptal flap (NSF) is sometimes used to prevent or repair postoperative cerebrospinal fluid (CSF) leaks. Few investigators have quantified long-term quality-of-life (QOL) outcomes regarding sinonasal measures after EETS, with or without involvement of the NSF. This study assesses whether the septal flap affects sinonasal QOL outcomes for patients receiving EETS for pituitary adenoma. METHODS AND MATERIALS: This is a retrospective study of patients who underwent EETS between 2013 and 2018. A total of 62 adults completed the Sinonasal Outcome Test-22 (SNOT-22) at least one year after the surgery. Outcome measures were compared between patients who underwent EETS with and without septal flap reconstruction. RESULTS: For the entire cohort, there were 14 patients (22.6%) who had septal flap reconstruction and 48 patients (77.4%) who did not. Patient demographics, tumor characteristics, surgical outcomes, and duration between surgery and completion of the questionnaire were similar for both groups. The mean SNOT-22 scores in the no reconstruction (NR) group and the nasoseptal flap reconstruction (NSFR) group were similar (P=0.9). In terms of SNOT-22 subdomains (rhinologic symptoms, extranasal rhinologic symptoms, ear/facial symptoms, psychological dysfunction, and sleep dysfunction), no significant differences were found when comparing the groups. CONCLUSION: As compared with no reconstructive involvement, NSF utilization does not affect the QOL and nasal symptoms of patients undergoing EETS.

Deuterium solid-state NMR quadrupolar order rotating frame relaxation with applications to amyloid-ß fibrils.

We describe a new method for measuring molecular dynamics based on the deuterium solid-state nuclear magnetic resonance (NMR) quadrupolar order rotating frame relaxation rate R1ρ,Q under static conditions. The observed quadrupolar order coherence is created using the broad-band Jeener-Broekaert excitation and is locked with a weak radio frequency (RF) field. We describe the experimental and theoretical approaches and show applications to a selectively deuterated valine side chain of the phosphorylated amyloid-ß (1-40) fibrils phosphorylated at the serine-8 position. The R1ρ,Q rate is sensitive to the rotameric exchange mode. For biological samples, the low spin-lock field in the 5- to 10-kHz range has the advantage of avoiding sample heating and dehydration. Thus, it provides an alternative to approaches based on single-quantum coherence, which require larger spin-lock fields.

Dynamics of Serine-8 Side-Chain in Amyloid-ß Fibrils and Fluorenylmethyloxycarbonyl Serine Amino Acid, Investigated by Solid-State Deuteron NMR.

Serine side-chains are strategic sites of post-translational modifications, and it is important to establish benchmarks of their internal dynamics. In this work, we compare the dynamics of serine side-chains in several biologically important systems: serine-8 in the disordered domain of Aß1-40 fibrils in the hydrated and dry states and fluorenylmethyloxycarbonyl (Fmoc) serine with the bulky group that mimics the hydrophobicity of the fibril contacts yet lacks the complexity of the protein system. Using deuterium solid-state NMR static line shape and longitudinal relaxation techniques in the 310 to 180 K temperature range, we compare the main features of the dynamics in these systems. The main motional modes in the fibrils are large-scale fluctuations in the hydrated state of the fibrils as well as local motions such as 3-site jumps of the Cß deuterons at high temperatures and small-angle fluctuations of the Cα-Cß axis at low temperatures. In the hydrated fibrils, two distinct states are present with vastly different extents of large-scale diffusive motions and 3-site-jump rate constants. The hydrated state at the physiological conditions is dominated by the "free" state undergoing large-scale diffusive motions and very fast local 3-site jumps, while in the "bound" state, these large-scale motions are quenched due to transient inter- and intramolecular interactions. Additionally, in the bound state, the 3-site-jump motions are orders of magnitude slower. Details of the dynamics in the serine side-chain are dependent on fine structural features and hydration levels of the systems.

Deuteron Quadrupolar Chemical Exchange Saturation Transfer (Q-CEST) Solid-State NMR for Static Powder Samples: Approach and Applications to Amyloid-ß Fibrils.

We provide an experimental and computational framework for 2 H quadrupolar chemical exchange saturation transfer NMR experiments (Q-CEST) under static solid-state conditions for the quantification of dynamics on µs-ms timescales. Simulations using simple 2-site exchange models provide insights into the relation between spin dynamics and motions. Biological applications focus on two sites of amyloid-ß fibrils in the 3-fold symmetric polymorph. The first site, the methyl group of A2 of the disordered N-terminal domain, undergoes diffusive motions and conformational exchange due to transient interactions. Earlier 2 H rotating frame relaxation and quadrupolar CPMG measurements are combined with the Q-CEST approach to characterize the multiple conformational states of the domain. The second site, the methyl group of M35, spans the water-accessible cavity inside the fibrils' core and undergoes extensive rotameric exchange. Q-CEST permits us to refine the rotameric exchange model for this site and allows the more precise determination of populations and rotameric exchange rate constants than line shape analysis.

Effect of Post-Translational Modifications and Mutations on Amyloid-ß Fibrils Dynamics at N Terminus.

We investigate the variability in the dynamics of the disordered N-terminal domain of amyloid-ß fibrils (Aß), comprising residues 1-16 of Aß1-40, due to post-translational modifications and mutations in the ß-bend regions known to modulate aggregation properties. Using 2H static solid-state NMR approaches, we compare the dynamics in the wild-type Aß fibrils in the threefold symmetric polymorph with the fibrils from three post-translational modification sequences: isoaspartate-D7, the phosphorylation of S8, and an N-terminal truncation ΔE3. Additional comparisons are made with the mutants in the ß-bend region (residues 21-23) corresponding to the familial Osaka E22Δ deletion and D23N Iowa mutation. We also include the aggregates induced by Zn2+ ions. The dynamics are probed at the F4 and G9 positions. The main motional model involves two free states undergoing diffusion and conformational exchanges with the bound state in which the diffusion is quenched because of transient interactions involving fibril core and other intrastrand contacts. The fraction of the bound state increases in a sigmoidal fashion with a decrease in temperature. There is clear variability in the dynamics: the phosphorylation of S8 variant is the most rigid at the G9 site in line with structural studies, the ΔE3 fibrils are more flexible at the G9 site in line with the morphological fragmentation pattern, the Zn-induced aggregates are the most mobile, and the two ß-bend mutants have the strongest changes at the F4 site toward higher rigidity. Overall, the changes underlie the potential role of conformational ensembles in setting the stage for aggregation-prone states.

Deuteron Solid-State NMR Relaxation Measurements Reveal Two Distinct Conformational Exchange Processes in the Disordered N-Terminal Domain of Amyloid-ß Fibrils.

We employed deuterium solid-state NMR techniques under static conditions to discern the details of the µs-ms timescale motions in the flexible N-terminal subdomain of Aß1-40 amyloid fibrils, which spans residues 1-16. In particular, we utilized a rotating frame (R1ρ ) and the newly developed time domain quadrupolar Carr-Purcell-Meiboom-Gill (QCPMG) relaxation measurements at the selectively deuterated side chains of A2, H6, and G9. The two experiments are complementary in terms of probing somewhat different timescales of motions, governed by the tensor parameters and the sampling window of the magnetization decay curves. The results indicated two mobile "free" states of the N-terminal domain undergoing global diffusive motions, with isotropic diffusion coefficients of 0.7-1 ⋅ 108 and 0.3-3 ⋅ 106 ad2 s-1 . The free states are also involved in the conformational exchange with a single bound state, in which the diffusive motions are quenched, likely due to transient interactions with the structured hydrophobic core. The conformational exchange rate constants are 2-3 ⋅ 105  s-1 and 2-3 ⋅ 104  s-1 for the fast and slow diffusion free states, respectively.