Biology subjects

Michaels, T. C. T.

Publications and source records attributed to Michaels, T. C. T..

12 recordsLinked to original sources

bioRxiv2026

Lipidic α-synuclein fibrils catalyze the formation of pathological mature fibrils

The aggregation of -synuclein into amyloid fibrils is a hallmark of a collection of neurodegenerative diseases termed synucleinopathies, including Parkinson's disease (PD). Interactions between -synuclein and lipid membranes modulate -synuclein aggregation and can produce protofibril structures consisting of both protein and lipid, termed lipidic fibrils. These lipidic fibrils can mature into mature fibrils, which resemble ex vivo fibrils extracted from Lewy bodies found in post-mortem brain samples of PD patients. Here, through the combination of experiments and kinetic theory, we study the influence of lipids and lipidic fibrils on the formation of mature fibrils. We find that lipidic fibrils catalyze mature fibril formation via a heterogeneous primary nucleation mechanism, which lowers the activation energy barrier of mature fibril formation relative to lipid-independent primary nucleation. The final stage of the reaction is governed by mature fibril elongation becoming limited by the rate at which protein monomers dissociate from the metastable lipidic fibrils, confirming that fibrils form in order of stability following Ostwald's Rule of Stages. This mechanism is validated using the small molecule inhibitor ambroxol, which selectively suppresses the formation of lipidic fibrils and in this manner shifts the aggregation kinetics to the slow, lipid-independent pathway. Taken together, these findings reveal that lipidic fibrils act as on-pathway intermediates and accelerate the aggregation of mature fibrils, which could provide a quantitative framework to design novel therapeutic strategies to inhibit pathological -synuclein aggregation.

biophysics↗

bioRxiv2026

Theory for Biomolecular Catalysis in Phase-Separated Systems

Enzymatic reactions in biomolecular condensates are often assumed to be regulated through local enrichment of reactants. However, condensates also reshape molecular transport and reaction kinetics, making it unclear how phase separation controls catalysis in living cells. Here, we develop a quantitative theory of biomolecular catalysis in phase-separated systems and find that liquid condensates can act as tunable catalytic switches, transitioning between regimes of enhanced and suppressed enzymatic activity, exhibiting optimal responses at biologically relevant condensate sizes. We show that condensate-mediated catalysis cannot be understood from reactant enrichment alone, but instead emerges from the coupled interplay of molecular partitioning, diffusive transport, and phase-dependent reaction kinetics. The strongest regulatory effects occur under rapid interphase exchange, where the spatially heterogeneous catalytic network admits a system-level Michaelis-Menten description governed by system-averaged concentrations and reaction kinetics. Our framework predicts that micron-sized condensates can either enhance or suppress enzymatic activity by up to two orders of magnitude, and that optimal catalytic regulation can emerge at condensate sizes comparable to many biomolecular condensates. These results provide experimentally testable predictions for condensate-mediated catalysis and establish quantitative principles for understanding and engineering enzyme-catalysed reactions in biomolecular condensates.

biophysics↗

bioRxiv2025

Self-consistent analytical solutions to the kinetics of lipid-induced protein aggregation

The aggregation of proteins into amyloid fibrils is a hallmark of several neurodegenerative disorders, including Parkinsons disease. A growing body of experimental evidence highlights the significant role lipid membranes play in modulating this aggregation process, particularly for proteins such as -synuclein. Despite this, there has been a lack of quantitative theoretical frameworks capable of describing the kinetics of lipid-induced protein aggregation. In this work, we develop an analytical model that explicitly incorporates lipid-mediated interactions into the aggregation kinetics. By formulating rate equations in terms of lipid surface coverage and applying a fixed-point analysis, we derive self-consistent solutions for the full timecourse of aggregation. Our model captures both one-step and two-step nucleation mechanisms and enables the prediction of key kinetic observables, including half-times and maximal growth rates. These results provide a quantitative foundation for interpreting experimental data and offer new mechanistic insights into how lipids influence the self-assembly of amyloidogenic proteins.

biophysics↗

bioRxiv2025

Structural defects in amyloid-β fibrils drive secondary nucleation

The nucleation of amyloid fibrils from monomeric protein, catalyzed by the surface of existing fibrils, is an important driver of many disorders such as Alzheimers and Parkinsons diseases. The structural basis of this secondary nucleation process, however, is poorly understood. Here, we ask whether secondary nucleation sites are found predominantly at rare growth defects: defects in the fibril core structure generated during their original assembly. We first demonstrate using the specific inhibitor of secondary nucleation, Brichos, that secondary nucleation sites on Alzheimers disease-associated fibrils composed of A{beta}40 and A{beta}42 peptides are rare compared to the number of protein molecules they contain. We then grow A{beta}40 fibrils under conditions designed to eliminate most growth defects while leaving the regular fibril morphology unchanged, and confirm the latter using cryo-electron microscopy. We measure both the ability of these annealed fibrils to promote secondary nucleation and the stoichiometry of their secondary nucleation sites, finding that both are greatly reduced as predicted. Re-analysis of published data for other proteins suggests that fibril growth defects that expose monomer planes or other structural units may also drive secondary nucleation generally, across most or all amyloids. These findings could unlock structure-based drug design of therapeutics that aim to halt amyloid disorders by inhibiting secondary nucleation sites.

biophysics↗

bioRxiv2024

Global kinetic model of lipid-induced alpha-synuclein aggregation and its inhibition by small molecules

The aggregation of -synuclein into amyloid fibrils is a hallmark of Parkinsons disease. This process has been shown to directly involve interactions between proteins and lipid surfaces when the latter are present. Despite this importance, the molecular mechanisms of lipid-induced amyloid aggregation have remained largely elusive. Here, we present a global kinetic model to describe lipid-induced amyloid aggregation of -synuclein. Using this framework we find that -synuclein fibrils form via a two-step primary nucleation mechanism and that lipid molecules are directly involved in both the nucleation and fibril elongation steps, giving rise to lipid-protein coaggregates. To illustrate the applicability of this kinetic approach to drug discovery, we identify the mechanism of action of squalamine, a known inhibitor of -synuclein aggregation, finding that this small molecule reduces the rate of lipid-dependent primary nucleation. Our work will likely guide the rational design of -synuclein aggregation inhibitors. Significance StatementAmyloid aggregation is a hallmark of a diverse range of diseases including Parkinsons Disease, where the protein -synuclein is a major con-stituent of proteinaceous deposits found in patients. It is well established that interactions between -synuclein and lipids modulate aggregation. However, the molecular mechanisms driving this lipid-induced aggregation have remained largely elusive, which has frustrated so far the discovery of drugs that prevent lipid-induced aggregation. In this work, we present the first global kinetic model describing lipid-induced aggregation and by integrating this theoretical framework with in vitro experimental data of lipid-induced -synuclein aggregation, we reveal the role of lipid membranes in the aggregation process and uncover the mechanism by which small molecule inhibitors interfere with this process.

biophysics↗

bioRxiv2023

Self-replication of Abeta42 aggregates occurs on small and isolated fibril sites

Self-replication of amyloid fibrils via secondary nucleation is an intriguing physicochemical phenomenon in which existing fibrils catalyse the formation of their own copies. The molecular events behind this fibril surface-mediated process remain largely inaccessible to current structural and imaging techniques. Using statistical mechanics, computer modelling, and chemical kinetics, we show that the catalytic structure of the fibril surface can be inferred from the aggregation behaviour in the presence and absence of a fibril-binding inhibitor. We apply our approach to the case of Alzheimers A{beta}42 amyloid fibrils formed in the presence of proSP-C Brichos inhibitors. We find that self-replication of A{beta}42 fibrils occurs on small catalytic sites on the fibril surface, which are far apart from each other, and each of which can be covered by a single Brichos inhibitor.

biophysics↗

bioRxiv2023

A solid beta-sheet structure is formed at the surface of FUS liquid droplets during aging

Insights into liquid droplet formation via liquid-liquid phase separation and the subsequent liquid-to-solid phase transition are important for understanding cell dynamics, as well as a number of neurodegenerative disorders. We report here, using the example of the FUsed in Sarcoma (FUS) protein, an investigation of the liquid droplet maturation process combining solution- and solid-state NMR spectroscopy, Raman spectroscopy, and light and electron microscopies. Our study reveals that the surface of the droplets plays a critical role in this process. Indeed, when comparing a biphasic sample, in which liquid droplets are stabilized in an agarose matrix, with a pure monophasic condensed phase sample, we find that the liquid-droplet maturation kinetics is faster in the biphasic FUS sample, owing to the larger surface-to-volume ratio. In addition, using Raman spectroscopy, we observe structural differences upon liquid-droplet maturation between the inside and the surface of liquid droplets, which is of {beta}-sheet content as revealed by solid-state NMR. This is detected very early on and increases over time. In agreement with these observations, a solid crust-like shell is visually seen by microaspiration experiments. After several months, electron microscopy reveals that the matured FUS droplets have converted into solid linear fibrils distinct from the fibril core of seeded fibrils reported previously, as arginine side-chains from the arginine-and-glycine-rich domain (RGG) motif are partially rigidified, highlighting the participation of this motif in the liquid-to-solid transition. In presence of RNA, this aging process is not taking place.

biochemistry↗

bioRxiv2023

The interplay between molecular assembly and phase separation

Many biological functions and dysfunctions rely on two fundamental processes, molecular assembly and the formation of condensed phases such as biomolecular condensates. Condensed phases generally form via phase separation, while molecular assemblies are clusters of molecules of various sizes, shapes, and functionality. We developed a theory that relies on thermodynamic principles to understand the interplay between molecular assembly and phase separation. We propose two prototypical classes of protein interactions and characterize their different equilibrium states and relaxation dynamics. We obtain results consistent with recent in vitro experimental observations of reconstituted proteins, including anomalous size distribution of assemblies, the gelation of condensed phases, and the change in condensate volume during ageing. Our theory provides the framework to unravel the mechanisms underlying physiological assemblies essential for cellular function, and aberrant assemblies which are associated with several neurodegenerative disorders.

biophysics↗

bioRxiv2022

Physical limits to acceleration of chemical reactions inside phase-separated compartments

We present a theoretical analysis of phase separated compartments as a means to facilitate chemical reactions. We find that the attractive interactions that concentrate reactants within the dense phase inhibit reactions by lowering the chemical potential and mobility of the reactants. Therefore, condensed phases are only beneficial if mobility in the condensed phase can be maintained. This can be achieved in multi-step reactions, where the proximity between enzymatic steps results in higher efficiency with less unreacted substrate, but does not increase the reaction rate. Alternatively, mobility can be maintained if recruitment to the condensed phase is driven by multiple attractive moieties that can bind and release independently. However, the spacers necessary to ensure independence between stickers are prone to entangle with the dense phase scaffold. We find an optimal sticker affinity that balances the need for rapid binding/unbinding kinetics and minimal entanglement. Reaction rates can be accelerated by shrinking the size of the dense phase with a corresponding increase in the number of stickers to enhance recruitment.

biophysics↗

bioRxiv2022

Self-consistent solution to the Flory-Huggins model and applications in protein liquid-liquid phase separation

A self-consistent analytical solution for binodal concentrations of the two-component Flory-Huggins model is derived. We show that this form extends the validity of the Ginzburg-Landau expansion away from the critical point to cover the whole phase space. Furthermore, this analytical solution reveals an exponential scaling law of the dilute phase binodal concentration as a function of interaction strength and chain length. We demonstrate explicitly the power of this approach by fitting experimental protein liquid-liquid phase separation boundaries to determine the effective chain length and the solute-solvent interaction energy. Moreover, we demonstrate that this strategy allows us to resolve the differences in the interaction energy contributions of individual amino acids. This analytical framework can serve as a new way to decode the protein sequence grammar for liquid-liquid phase separation.

biophysics↗

bioRxiv2021

Aggregation controlled by condensate rheology

Biomolecular condensates in living cells can exhibit a complex rheology including viscoelastic and glassy behaviour. This rheological behavior of condensates was suggested to regulate polymerisation of cytoskeletal filaments and aggregation of amyloid fibrils. Here, we theoretically investigate how the rheological properties of condensates can control the formation of linear aggregates. To this end, we propose a kinetic theory for linear aggregation in coexisting phases, which accounts for the aggregate size distribution and the exchange of aggregates between inside and outside of condensates. The rheology of condensates is accounted in our model via aggregate mobilities that depend on aggregate size. We show that condensate rheology determines whether aggregates of all sizes or dominantly small aggregates are exchanged between condensate inside and outside on the time-scale of aggregation. As a result, the ratio of aggregate numbers inside to outside of condensates differs significantly. Strikingly, we also find that weak variations in the rheological properties of condensates can lead to a switch-like change of the number of aggregates. These results suggest a possible physical mechanism for how living cells could control linear aggregation in a switch-like fashion through variations in condensate rheology. SIGNIFICANCEThe intracellular space can be organized through phase-separated condensates that often exhibit rheological properties reminiscent of complex fluids. These condensates can affect biochemical processes such as the formation of linear aggregates, in particular biofilaments or amyloids. Here, we propose a theoretical model for how condensate rheology can control the irreversible formation of linear aggregates. A key finding is that size and number of aggregates change in a switch-like fashion upon weak changes in condensate rheology. Our model paves the way to unravel the physiochemical mechanisms of how the rheology of condensates can control aberrant protein aggregation. Such mechanisms may explain how rheological changes, such as ageing or the transition to dormancy, give rise to diseases related to protein aggregation.

biophysics↗

bioRxiv2020

Kinetic analysis reveals the rates and mechanisms of protein aggregation in a multicellular organism

The accumulation of insoluble protein aggregates containing amyloid fibrils has been observed in many different human protein misfolding diseases1,2, and their pathological features have been recapitulated in diverse model systems3. In vitro kinetic studies have provided a quantitative understanding of how the fundamental molecular level processes of nucleation and growth lead to amyloid formation4. However, it is not yet clear to what extent these basic biophysical processes translate to amyloid formation in vivo, given the complexity of the cellular and organismal environment. Here we show that the aggregation of a fluorescently tagged polyglutamine (polyQ) protein into {micro}m-sized inclusions in the muscle tissue of living C. elegans can be quantitatively described by a molecular model where stochastic nucleation occurs independently in each cell, followed by rapid aggregate growth. Global fitting of the image-based aggregation kinetics reveals a nucleation rate corresponding to 0.01 h-1 per cell at 1 mM intracellular protein concentration, and shows that the intrinsic stochasticity of nucleation accounts for a significant fraction of the observed animal-to-animal variation. Our results are consistent with observations for the aggregation of polyQ proteins in vitro5 and in cell culture6, and highlight how nucleation events control the overall progression of aggregation in the organism through the spatial confinement into individual cells. The key finding that the biophysical principles associated with protein aggregation in small volumes remain the governing factors, even in the complex environment of a living organism, will be critical for the interpretation of in vivo data from a wide range of protein aggregation diseases.

biophysics↗