RESUMO
Metal ions, including biologically prevalent sodium ions, can modulate electrostatic interactions frequently involved in the stability of condensed compartments in cells. Quantitative characterization of heterogeneous ion dynamics inside biomolecular condensates demands new experimental approaches. Here we develop a 23Na NMR relaxation-based integrative approach to probe dynamics of sodium ions inside agarose gels as a model system. We exploit the electric quadrupole moment of spin-3/2 23Na nuclei and, through combination of single-quantum and triple-quantum-filtered 23Na NMR relaxation methods, disentangle the relaxation contribution of different populations of sodium ions inside gels. Three populations of sodium ions are identified: a population with bi-exponential relaxation representing ions within the slow motion regime and two populations with mono-exponential relaxation but at different rates. Our study demonstrates the dynamical heterogeneity of sodium ions inside agarose gels and presents a new experimental approach for monitoring dynamics of sodium and other spin-3/2 ions (e.g. chloride) in condensed environments.
Assuntos
Géis , Sefarose , Sódio , Sefarose/química , Sódio/química , Géis/química , Espectroscopia de Ressonância Magnética/métodos , Íons/química , Teoria QuânticaRESUMO
Signal transduction by the ligated B cell antigen receptor (BCR) depends on the preorganization of its intracellular components, such as the effector proteins SLP65 and CIN85 within phase-separated condensates. These liquid-like condensates are based on the interaction between three Src homology 3 (SH3) domains and the corresponding proline-rich recognition motifs (PRM) in CIN85 and SLP65, respectively. However, detailed information on the protein conformation and how it impacts the capability of SLP65/CIN85 condensates to orchestrate BCR signal transduction is still lacking. This study identifies a hitherto unknown intramolecular SH3:PRM interaction between the C-terminal SH3 domain (SH3C) of CIN85 and an adjacent PRM. We used high-resolution nuclear magnetic resonance (NMR) experiments to study the flexible linker region containing the PRM and determined the extent of the interaction in multidomain constructs of the protein. Moreover, we observed that the phosphorylation of a serine residue located in the immediate vicinity of the PRM regulates this intramolecular interaction. This allows for a dynamic modulation of CIN85's valency toward SLP65. B cell culture experiments further revealed that the PRM/SH3C interaction is crucial for maintaining the physiological level of SLP65/CIN85 condensate formation, activation-induced membrane recruitment of CIN85, and subsequent mobilization of Ca2+. Our findings therefore suggest that the intramolecular interaction with the adjacent disordered linker is effective in modulating CIN85's valency both in vitro and in vivo. This therefore constitutes a powerful way for the modulation of SLP65/CIN85 condensate formation and subsequent B cell signaling processes within the cell.
Assuntos
Proteínas Adaptadoras de Transdução de Sinal , Transdução de Sinais , Proteínas Adaptadoras de Transdução de Sinal/química , Transdução de Sinais/fisiologia , Domínios de Homologia de src , Linfócitos B/metabolismo , Receptores de Antígenos de Linfócitos B/metabolismo , Ligação ProteicaRESUMO
Alzheimer's disease is characterized by the presence of extraneuronal amyloid plaques composed of amyloid-beta (Aß) fibrillar aggregates in the brains of patients. In mouse models, it has previously been shown that atorvastatin (Ator), a cholesterol-lowering drug, has some reducing effect on the production of cerebral Aß. A meta-analysis on humans showed moderate effects in the short term but no improvement in the Alzheimer's Disease Assessment Scale-Cognitive Subscale behavioral test. Here, we explore a potential direct effect of Ator on Aß42 aggregation. Using NMR-based monomer consumption assays and CD spectroscopy, we observed a promoting effect of Ator in its original form (Ator-calcium) on Aß42 aggregation, as expected because of the presence of calcium ions. The effect was reversed when applying a CaCO3-based calcium ion scavenging method, which was validated by the aforementioned methods as well as thioflavin-T fluorescence assays and transmission electron microscopy. We found that the aggregation was inhibited significantly when the concentration of calcium-free Ator exceeded that of Aß by at least a factor of 2. The 1H-15N heteronuclear single quantum correlation and saturation-transfer difference NMR data suggest that calcium-free Ator exerts its effect through interaction with the 16KLVF19 binding site on the Aß peptide via its aromatic rings as well as hydroxyl and methyl groups. On the other hand, molecular dynamics simulations confirmed that the increasing concentration of Ator is necessary for the inhibition of the conformational transition of Aß from an α-helix-dominant to a ß-sheet-dominant structure.
Assuntos
Doença de Alzheimer , Peptídeos beta-Amiloides , Atorvastatina , Fragmentos de Peptídeos , Doença de Alzheimer/tratamento farmacológico , Doença de Alzheimer/metabolismo , Amiloide/metabolismo , Peptídeos beta-Amiloides/metabolismo , Animais , Atorvastatina/farmacologia , Cálcio/metabolismo , Humanos , Camundongos , Fragmentos de Peptídeos/metabolismoRESUMO
Reorientational dynamics of intrinsically disordered proteins (IDPs) contain multiple motions often clustered around three motional modes: ultrafast librational motions of amide groups, fast local backbone conformational fluctuations and slow chain segmental motions. This dynamic picture is mainly based on 15 Nâ NMR relaxation studies of IDPs at relatively low temperatures where the amide-water proton exchange rates are sufficiently small. Less is known, however, about the dynamics of IDPs at more physiological temperatures. Here, we investigate protein dynamics in a 441-residue long IDP, tau protein, in the temperature range from 0-25 °C, using 15 Nâ NMR relaxation rates and spectral density analysis. While at these temperatures relaxation rates are still better described in terms of amide group librational motions, local backbone dynamics and chain segmental motions, the temperature-dependent trend of spectral densities suggests that the timescales of fast backbone conformational fluctuations and slower chain segmental motions might become inseparable at higher temperatures. Our data demonstrate the remarkable dynamic plasticity of this prototypical IDP and highlight the need for dynamic studies of IDPs at multiple temperatures.
Assuntos
Proteínas Intrinsicamente Desordenadas , Proteínas tau , Temperatura , Conformação Proteica , Espectroscopia de Ressonância Magnética , Proteínas Intrinsicamente Desordenadas/química , AmidasRESUMO
Liquid water is essential for life as we know it and the coupling between water and biomolecular dynamics is crucial for life processes. Jupiter's moon Europa is a good candidate for searching for extraterrestrial life in our outer solar system, mainly because a liquid water salty ocean in contact with a rocky seafloor underlies its ice shell. Little, however, is known about the chemical composition of the subglacial ocean of Europa or the brine pockets within its ice shell and their impacts on water dynamics. Here, we employ 1H, 17O, 23Na and 35Cl NMR spectroscopy, especially NMR spin relaxation and diffusion methods, and investigate the mobility of water molecules and ions in eutectic solutions of magnesium sulfate and sodium chloride, two salts ubiquitously present on the surface of Europa, over a range of temperatures and pressures pertinent to Europa's subglacial ocean. The NMR data demonstrate the more pronounced effect of magnesium sulfate compared with sodium chloride on the mobility of water molecules. Even at its much lower eutectic temperature, the sodium chloride solution retains a relatively large level of water mobility. Our results highlight the higher potential of a sodium chloride-rich than magnesium sulfate-rich Europa's ocean to accommodate life and support life origination within the eutectic melts of Europa's ice shell.
RESUMO
Little is known about how maturation of Alzheimer's disease-related amyloid ß (Aß) fibrils alters their stability and potentially influences their spreading in the brain. Using high-pressure NMR, we show that progression from early to late Aß40 aggregates enhances the kinetic stability, while ageing during weeks to months enhances their thermodynamic stability.
Assuntos
Doença de Alzheimer , Peptídeos beta-Amiloides , Humanos , Peptídeos beta-Amiloides/química , Espectroscopia de Ressonância Magnética , Amiloide/química , Fragmentos de Peptídeos/químicaRESUMO
Micrometer-sized objects are widely known to exhibit chemically driven motility in systems away from equilibrium. Experimental observation of reaction-induced motility or enhancement in diffusivity at the much shorter length scale of small molecules is, however, still a matter of debate. Here, we investigate the molecular diffusivity of reactants, catalyst, and product of a model reaction, the copper-catalyzed azide-alkyne cycloaddition click reaction, and develop new NMR diffusion approaches that allow the probing of reaction-induced diffusion enhancement in nanosized molecular systems with higher accuracy than the state of the art. Following two different approaches that enable the accounting of time-dependent concentration changes during NMR experiments, we closely monitored the diffusion coefficient of reaction components during the reaction. The reaction components showed distinct changes in the diffusivity: while the two reactants underwent a time-dependent decrease in their diffusivity, the diffusion coefficient of the product gradually increased and the catalyst showed only slight diffusion enhancement within the range expected for reaction-induced sample heating. The decrease in diffusion coefficient of the alkyne, one of the two reactants of click reaction, was not reproduced during its copper coordination when the second reactant, azide, was absent. Our results do not support the catalysis-induced diffusion enhancement of the components of the click reaction and, instead, point to the role of a relatively large intermediate species within the reaction cycle with diffusivity lower than that of both the reactants and product molecule.
RESUMO
In their Comment (DOI: 10.1021/jacs.2c02965) on two related publications by our group (J. Am. Chem. Soc. 2022, 144, 1380-1388; DOI: 10.1021/jacs.1c11754) and another (J. Am. Chem. Soc. 2021, 143, 20884-20890; DOI: 10.1021/jacs.1c09455), Huang and Granick refer to the diffusion NMR measurements of molecules during a copper-catalyzed azide-alkyne cycloaddition (CuAAC) "click" reaction. Here we respond to their comments and maintain that no measurable diffusion enhancement was observed during the reaction. We expand on the physical arguments presented in our original JACS Article regarding the appropriate reference state for the diffusion coefficient and present new data showing that the use of other reference states, as suggested by Huang and Granick, will still support our conclusion that the two reactants and one product of the CuAAC reaction do not exhibit boosted mobility during the reaction.
Assuntos
Azidas , Química Click , Alcinos/química , Azidas/química , Catálise , Cobre/química , Reação de CicloadiçãoRESUMO
Biomolecular phase separation plays a key role in the spatial organization of cellular activities. Dynamic formation and rapid component exchange between phase separated cellular bodies and their environment are crucial for their function. Here, we employ a well-established phase separating model system, namely, a triethylamine (TEA)-water mixture, and develop an NMR approach to detect the exchange of scaffolding TEA molecules between separate phases and determine the underlying exchange rate. We further demonstrate how the advantageous NMR properties of fluorine nuclei provide access to otherwise inaccessible exchange processes of a client molecule. The developed NMR-based approach allows quantitative monitoring of the effect of regulatory factors on component exchange and facilitates "exchange"-based screening and optimization of small molecules against druggable biomolecular targets located inside condensed phases.
Assuntos
Flúor , Imageamento por Ressonância Magnética , Humanos , Espectroscopia de Ressonância Magnética , Modelos Biológicos , Ressonância Magnética Nuclear BiomolecularRESUMO
Heparins and heparan sulfate polysaccharides are negatively charged glycosaminoglycans and play important roles in cell-to-matrix and cell-to-cell signaling processes. Metal ion binding to heparins alters the conformation of heparins and influences their function. Various experimental techniques have been used to investigate metal ion-heparin interactions, frequently with inconsistent results. Exploiting the quadrupolar 23Na nucleus, we herein develop a 23Na NMR-based competition assay and monitor the binding of divalent Ca2+ and Mg2+ and trivalent Al3+ metal ions to sodium heparin and the consequent release of sodium ions from heparin. The 23Na spin relaxation rates and translational diffusion coefficients are utilized to quantify the metal ion-induced release of sodium ions from heparin. In the case of the Al3+ ion, the complementary approach of 27Al quadrupolar NMR is employed as a direct probe of ion binding to heparin. Our NMR results demonstrate at least two metal ion-binding sites with different affinities on heparin, potentially undergoing dynamic exchange. For the site with lower metal ion binding affinity, the order of Ca2+ > Mg2+ > Al3+ is obtained, in which even the weakly binding Al3+ ion is capable of displacing sodium ions from heparin. Overall, the multinuclear quadrupolar NMR approach employed here can monitor and quantify metal ion binding to heparin and capture different modes of metal ion-heparin binding.
Assuntos
Heparina , Heparitina Sulfato , Heparina/química , Espectroscopia de Ressonância Magnética/métodos , Heparitina Sulfato/metabolismo , Metais/metabolismo , Íons , Sódio/metabolismo , Sítios de LigaçãoRESUMO
The amyloid cascade hypothesis proposes that amyloid-beta (Aß) aggregation is the initial triggering event in Alzheimer's disease. Here, we utilize NMR spectroscopy and monitor the structural dynamics of two variants of Aß, Aß40 and Aß42, as a function of temperature. Despite having identical amino acid sequence except for the two additional C-terminal residues, Aß42 has higher aggregation propensity than Aß40. As revealed by the NMR data on dynamics, including backbone chemical shifts, intra-methyl cross-correlated relaxation rates and glycine-based singlet-states, the C-terminal region of Aß, especially the G33-L34-M35 segment, plays a particular role in the early steps of temperature-induced Aß aggregation. In Aß42, the distinct dynamical behaviour of C-terminal residues at higher temperatures is accompanied with marked changes in the backbone dynamics of residues V24-K28. The distinctive role of the C-terminal region of Aß42 in the initiation of aggregation defines a target for the rational design of Aß42 aggregation inhibitors.
Assuntos
Peptídeos beta-Amiloides/química , Humanos , Ressonância Magnética Nuclear Biomolecular , Conformação Proteica , Dobramento de ProteínaRESUMO
Intrinsically disordered proteins (IDPs) experience a diverse spectrum of motions that are difficult to characterize with a single experimental technique. Herein we combine high- and low-field nuclear spin relaxation, nanosecond fluorescence correlation spectroscopy (nsFCS), and long molecular dynamics simulations of alpha-synuclein, an IDP involved in Parkinson disease, to obtain a comprehensive picture of its conformational dynamics. The combined analysis shows that fast motions below 2â ns caused by local dihedral angle fluctuations and conformational sampling within and between Ramachandran substates decorrelate most of the backbone N-H orientational memory. However, slow motions with correlation times of up to ca. 13â ns from segmental dynamics are present throughout the alpha-synuclein chain, in particular in its C-terminal domain, and global chain reconfiguration occurs on a timescale of ca. 60â ns. Our study demonstrates a powerful strategy to determine residue-specific protein dynamics in IDPs at different time and length scales.
Assuntos
Proteínas Intrinsicamente Desordenadas/química , alfa-Sinucleína/química , Humanos , Simulação de Dinâmica Molecular , Doença de Parkinson/patologia , Agregação Patológica de Proteínas/patologia , Conformação Proteica , Domínios Proteicos , Dobramento de Proteína , Espectrometria de FluorescênciaRESUMO
Neurodegeneration is characterized by the ubiquitous presence of modifications in protein deposits. Despite their potential significance in the initiation and progression of neurodegenerative diseases, the effects of posttranslational modifications on the molecular properties of protein aggregates are largely unknown. Here, we study the Alzheimer disease-related amyloid-ß (Aß) peptide and investigate how phosphorylation at serine 8 affects the structure of Aß aggregates. Serine 8 is shown to be located in a region of high conformational flexibility in monomeric Aß, which upon phosphorylation undergoes changes in local conformational dynamics. Using hydrogen-deuterium exchange NMR and fluorescence quenching techniques, we demonstrate that Aß phosphorylation at serine 8 causes structural changes in the N-terminal region of Aß aggregates in favor of less compact conformations. Structural changes induced by serine 8 phosphorylation can provide a mechanistic link between phosphorylation and other biological events that involve the N-terminal region of Aß aggregates. Our data therefore support an important role of posttranslational modifications in the structural polymorphism of amyloid aggregates and their modulatory effect on neurodegeneration.
Assuntos
Peptídeos beta-Amiloides/química , Agregação Patológica de Proteínas , Doença de Alzheimer/metabolismo , Peptídeos beta-Amiloides/metabolismo , Medição da Troca de Deutério , Humanos , Ressonância Magnética Nuclear Biomolecular , Fosforilação , Serina/química , Serina/metabolismoRESUMO
Molecular dynamics (MD) at two temperatures of 300 and 340 K identified two histidine residues, His461 and His489, in the most flexible regions of firefly luciferase, a light emitting enzyme. We therefore designed four protein mutants H461D, H489K, H489D and H489M to investigate their enzyme kinetic and thermodynamic stability changes. Substitution of His461 by aspartate (H461D) decreased ATP binding affinity, reduced the melting temperature of protein by around 25 °C and shifted its optimum temperature of activity to 10 °C. In line with the common feature of psychrophilic enzymes, the MD data showed that the overall flexibility of H461D was relatively high at low temperature, probably due to a decrease in the number of salt bridges around the mutation site. On the other hand, substitution of His489 by aspartate (H489D) introduced a new salt bridge between the C-terminal and N-terminal domains and increased protein rigidity but only slightly improved its thermal stability. Similar changes were observed for H489K and, to a lesser degree, H489M mutations. Based on our results we conclude that the MD simulation-based rational substitution of histidines by salt-bridge forming residues can modulate conformational dynamics in luciferase and shift its optimal temperature activity.
Assuntos
Substituição de Aminoácidos , Histidina , Luciferases de Vaga-Lume/química , Luciferases de Vaga-Lume/metabolismo , Temperatura , Sequência de Aminoácidos , Sequência de Bases , Estabilidade Enzimática/genética , Ligação de Hidrogênio , Cinética , Luciferases de Vaga-Lume/genética , Simulação de Dinâmica Molecular , Mutação , Conformação ProteicaRESUMO
Biological control systems regulate the behavior of biological systems in a constantly changing environment. Homeostasis is the most widely studied outcome of biological control systems. Homeostatic systems maintain the system in its desired state despite variations in system parameters or the externally-determined input rates of their constituents, i.e. they have zero or near zero steady state error. On the other hand, allostatic systems are not resistant against environmental changes and the steady state level of their controlled variables responds positively to the changes in their input rates. Little is known, however, on the existence and frequency of reverse allostatic systems, where the steady state value of the controlled variable correlates negatively with the input rate of that variable. In the present study, we derive the minimal conditions for the existence and local stability of reverse allostatic systems, and demonstrate in examples of metabolic, pharmacological, pathophysiological and ecological systems that the reverse allostasis requirements are relatively non-stringent and may be satisfied in biological systems more commonly than usually thought. The possible existence of reverse allostatic systems in nature and their counter-intuitive implications in physiological systems, drug treatment, ecosystem management, and biological control are explored and testable predictions are made.
Assuntos
Alostase/fisiologia , Modelos Biológicos , Adaptação Fisiológica , Animais , Homeostase , Humanos , Estresse FisiológicoRESUMO
MOTIVATION: A large fraction of eukaryotic proteins contain unstructured tails or linkers. The presence of flexible regions allows these systems to experience a high level of mobility facilitating their biological function. The complex nature of protein rotation in such flexible modular systems precludes a straightforward application of hydrodynamic methods to calculate their rotational motional properties. We describe the workflow of HYdrodynamic CoUpling of Domains (HYCUD), a program for prediction of effective rotational correlation times in multidomain proteins. The usage of HYCUD is demonstrated by its application to the ribosomal protein L7/L12. Rotational correlation times predicted by HYCUD might be used to detect molecular switch events mediated by disorder-order transitions in interdomain linkers. AVAILABILITY AND IMPLEMENTATION: The source code and documentation are available at www.mpibpc.mpg.de/106144/software. CONTACT: mzwecks@gwdg.de or nare@nmr.mpibpc.mpg.de SUPPLEMENTARY INFORMATION: Supplementary material is available at Bioinformatics online.
Assuntos
Biologia Computacional/métodos , Proteínas Ribossômicas/química , Rotação , Software , Estrutura Terciária de ProteínaRESUMO
Aggregation and toxicity of the amyloid ß-peptide (Aß) are considered as critical events in the initiation and progression of Alzheimer's disease (AD). Recent evidence indicated that soluble oligomeric Aß assemblies exert pronounced toxicity, rather than larger fibrillar aggregates that deposit in the forms of extracellular plaques. While some rare mutations in the Aß sequence that cause early-onset AD promote the oligomerization, molecular mechanisms that induce the formation or stabilization of oligomers of the wild-type Aß remain unclear. Here, we identified an Aß variant phosphorylated at Ser26 residue (pSer26Aß) in transgenic mouse models of AD and in human brain that shows contrasting spatio-temporal distribution as compared to non-phosphorylated Aß (npAß) or other modified Aß species. pSer26Aß is particularly abundant in intraneuronal deposits at very early stages of AD, but much less in extracellular plaques. pSer26Aß assembles into a specific oligomeric form that does not proceed further into larger fibrillar aggregates, and accumulates in characteristic intracellular compartments of granulovacuolar degeneration together with TDP-43 and phosphorylated tau. Importantly, pSer26Aß oligomers exert increased toxicity in human neurons as compared to other known Aß species. Thus, pSer26Aß could represent a critical species in the neurodegeneration during AD pathogenesis.
Assuntos
Doença de Alzheimer/metabolismo , Doença de Alzheimer/patologia , Peptídeos beta-Amiloides/metabolismo , Encéfalo/metabolismo , Serina/metabolismo , Fatores Etários , Doença de Alzheimer/genética , Peptídeos beta-Amiloides/toxicidade , Precursor de Proteína beta-Amiloide/genética , Animais , Diferenciação Celular , Células Cultivadas , Modelos Animais de Doenças , Regulação da Expressão Gênica/genética , Humanos , Camundongos , Camundongos Transgênicos , Células-Tronco Neurais , Neuroblastoma/patologia , Fosforilação/genética , Agregados Proteicos/genética , Fatores de Tempo , TransfecçãoRESUMO
In addition to the prototypic amyloid-ß (Aß) peptides Aß1-40 and Aß1-42 , several Aß variants differing in their amino and carboxy termini have been described. Synthetic availability of an Aß variant is often the key to study its role under physiological or pathological conditions. Herein, we report a protocol for the efficient solid-phase peptide synthesis of the N-terminally elongated Aß-peptides Aß-3-38 , Aß-3-40 , and Aß-3-42 . Biophysical characterization by NMR spectroscopy, CD spectroscopy, an aggregation assay, and electron microscopy revealed that all three peptides were prone to aggregation into amyloid fibrils. Immunoprecipitation, followed by mass spectrometry, indicated that Aß-3-38 and Aß-3-40 are generated by transfected cells even in the presence of a tripartite ß-site amyloid precursor protein cleaving enzymeâ 1 (BACE1) inhibitor. The elongated Aß peptides starting at Val(-3) can be separated from N-terminally-truncated Aß forms by high-resolution isoelectric-focusing techniques, despite virtually identical isoelectric points. The synthetic Aß variants and the methods presented here are providing tools to advance our understanding of the potential roles of N-terminally elongated Aß variants in Alzheimer's disease.
Assuntos
Peptídeos beta-Amiloides/síntese química , Sequência de Aminoácidos , Peptídeos beta-Amiloides/química , Peptídeos beta-Amiloides/metabolismo , Benzotiazóis , Dicroísmo Circular , Humanos , Cinética , Espectrometria de Massas , Microscopia Eletrônica , Ressonância Magnética Nuclear Biomolecular , Técnicas de Síntese em Fase Sólida , Tiazóis/química , Tiazóis/metabolismoRESUMO
Alzheimer's disease (AD) is the most common form of dementia and associated with progressive deposition of amyloid ß-peptides (Aß) in the brain. Aß derives by sequential proteolytic processing of the amyloid precursor protein by ß- and γ-secretases. Rare mutations that lead to amino-acid substitutions within or close to the Aß domain promote the formation of neurotoxic Aß assemblies and can cause early-onset AD. However, mechanisms that increase the aggregation of wild-type Aß and cause the much more common sporadic forms of AD are largely unknown. Here, we show that extracellular Aß undergoes phosphorylation by protein kinases at the cell surface and in cerebrospinal fluid of the human brain. Phosphorylation of serine residue 8 promotes formation of oligomeric Aß assemblies that represent nuclei for fibrillization. Phosphorylated Aß was detected in the brains of transgenic mice and human AD brains and showed increased toxicity in Drosophila models as compared with non-phosphorylated Aß. Phosphorylation of Aß could represent an important molecular mechanism in the pathogenesis of the most common sporadic form of AD.
Assuntos
Doença de Alzheimer/patologia , Peptídeos beta-Amiloides/metabolismo , Desnaturação Proteica , Animais , Encéfalo/patologia , Modelos Animais de Doenças , Drosophila , Humanos , Camundongos , Camundongos Transgênicos , Modelos Biológicos , FosforilaçãoRESUMO
Pathogenesis of Alzheimer's disease (AD) is associated with aggregation of the amyloid-ß (Aß) peptide into oligomeric and fibrillar assemblies; however, little is known about the molecular basis of aggregation of Aß into distinct assembly states. Here we demonstrate that phosphorylation at serine 26 (S26) impairs Aß fibrillization while stabilizing its monomers and nontoxic soluble assemblies of nonfibrillar morphology. NMR spectroscopy and replica-exchange molecular dynamics indicate that introduction of a phosphate group or phosphomimetic at position 26 diminishes Aß's propensity to form a ß-hairpin, rigidifies the region around the modification site, and interferes with formation of a fibril-specific salt bridge between aspartic acid 23 and lysine 28. The combined data demonstrate that phosphorylation of S26 prevents a distinct conformational rearrangement that is required for progression of Aß aggregation toward fibrils and provide a basis for a possible role of phosphorylation at serine 26 in AD.