Biology subjects

Toprakcioglu, Z.

Publications and source records attributed to Toprakcioglu, Z..

10 recordsLinked to original sources

bioRxiv2026

Melanin Suppresses Aβ Aggregation and Toxicity

The aggregation of amyloid-{beta} (A{beta}) peptides into insoluble deposits is a characteristic hallmark of Alzheimer's disease (AD) and related neurodegenerative disorders. While AD is the most common cause of dementia, there are currently no disease-modifying treatments which are both affordable and adverse-free. In this study, we report that melanin, a pigment which is commonly found in nature and is abundant in parts of the human brain, suppresses the aggregation of the 42-amino acid A{beta} variant (A{beta}42). Using biophysical and biochemical techniques, we show that melanin delays A{beta}42 aggregation while also reducing the amount of A{beta}42 that converts into aggregates. Using thioflavin T assays paired with chemical kinetics, we characterised the melanin-induced inhibition of A{beta}42 aggregation in vitro. Using MALDI-MS, we elucidate the molecular basis of this effect by showing that melanin prevents A{beta}42 dimerisation. We then demonstrate that melanin also reverts the aggregation process by dissolving pre-formed A{beta}42 fibrils. Finally, we show that melanin reduces A{beta}42 aggregation and rescues A{beta}42 toxicity in an SH-SY5Y neuroblastoma cell model. Our study shows that melanin disrupts the aggregation and cytotoxicity of A{beta}42, and suggests that compounds derived from human metabolites may offer promising avenues to combat amyloid formation.

biophysics↗

bioRxiv2026

Low-abundance αSyn-112 promotes αSyn-140 aggregation in vitro and forms immunoreactive deposits in Parkinson's disease brain tissue

The aggregation of -synuclein (Syn) is a molecular hallmark of Parkinsons disease (PD) and other synucleinopathies. Understanding the molecular mechanisms that determine the aggregation of this protein may thus facilitate the development of disease-modifying therapies. While Syn is most commonly expressed as a 140-residue protein (Syn-140), recent evidence suggests an involvement of alternatively spliced Syn isoforms in disease onset and progression. Here, we report and characterise the interaction between Syn-140 and the aggregation-prone Syn-112 variant, one of the most abundant Syn splice isoforms. We found that amounts as low as 1% of Syn-112 accelerate the nucleation and aggregation of Syn-140. To further investigate this phenomenon, we employed MALDI-MS and NMR spectroscopy, confirming that Syn-140 and Syn-112 monomers interact strongly with one another. Furthermore, to assess the association of Syn-112 with disease pathology, we performed immunohistochemical staining combined with confocal microscopy on PD brain samples. Thereby, we found an increase in the number as well as the area of Syn-112 immunoreactive aggregates compared to healthy controls. These results illustrate how low-abundance Syn splice isoforms can modulate the aggregation landscape of Syn-140 and in turn contribute to the molecular heterogeneity of synucleinopathies.

biophysics↗

bioRxiv2026

Nuclear Pct1 couples phosphatidylcholine synthesis with membrane biogenesis

Phosphatidylcholine (PC) is the major eukaryotic phospholipid and its synthesis must be homeostatically controlled to prevent excess membrane and organelle growth. Here we investigate how PC synthesis by the Kennedy pathway is coordinated with membrane biogenesis. In budding yeast, the rate-limiting enzyme Pct1 is nuclear and reversibly associates with the inner nuclear membrane (INM) in response to lipid packing defects caused by low PC. We show that the enzymes acting after Pct1 to generate PC remain at the endoplasmic reticulum (ER) during pathway activation. Relocating the final PC synthesis step to different endomembrane sites does not alter the kinetics of Pct1 release from the INM, indicating that newly made PC equilibrates with the INM rapidly. In contrast, elevated phosphatidic acid locks Pct1 at the INM, prevents pathway inactivation and drives nuclear/ER membrane proliferation. These results support a model in which nuclear Pct1 senses lipid imbalance while ER-localized enzymes supply PC; disrupting this homeostasis leads to uncontrolled membrane biogenesis.

cell biology↗

bioRxiv2023

Nanoscale profiling of evolving intermolecular interactions in ageing FUS condensates

In addition to the native state, proteins can form liquid-like condensates, viscoelastic condensates, such as gels, as well as solid-like condensates, such as amyloid fibrils, crystals and amorphous materials. The material properties of these condensates play important roles in their cellular functions, with aberrant liquid-to-solid phase transitions having been implicated in neurodegenerative diseases. However, the molecular changes and resultant material properties across the whole phase space of condensates are complex and yet to be fully understood. The extreme sensitivity to their environment, which enables their biological function, is also what makes protein condensates particularly challenging experimental targets. Here, we provide a characterisation of the ageing behaviour of the full-length fused in sarcoma (FUS) protein. We achieve this goal by using a microfluidic sample deposition technology to enable the application of surface-based techniques to the study of biological condensates. We first demonstrate that we maintain relevant structural features of condensates in physiologically-relevant conditions on surfaces. Then, using a combination of atomic force microscopy and vibrational spectroscopy, we characterise the spatio-temporal changes in the structure and mechanical properties of the condensates to reveal local phase transitions in individual condensates. We observe that initially dynamic, fluid-like condensates undergo a global increase in elastic response conferred by an increase in the density of cation-{pi} intermolecular interactions. Solid-like structures form first at condensate-solvent interfaces, before heterogeneously propagating throughout the aged fluid core. These solid structures are composed of heterogenous, non-amyloid {beta}-sheets, which are stabilised by hydrogen-bonding interactions not observed in the fluid state. Overall, this study identifies the molecular conformations associated with different physical states of FUS condensates, establishing a technology platform to understand the role of phase behaviour in condensate function and dysfunction.

biophysics↗

bioRxiv2023

Aggregation of the Amyloid-beta Peptide (Abeta40) within Condensates Generated through Liquid-Liquid Phase Separation

The deposition of the A{beta} peptide into amyloid fibrils is characteristic of Alzheimers disease. As it has been recently observed that the process of amyloid aggregation can take place within an intermediate liquid-like condensed phase, we investigated whether A{beta} could undergo liquid-liquid phase separation, and whether A{beta} amyloid aggregation could take place within A{beta} liquid condensates. By using a microfluidic protocol, we observed that the 40-residue form of A{beta} (A{beta}40) can undergo liquid-liquid phase separation, and that accessing a liquid intermediate state enhances primary nucleation and enables A{beta}40 to readily self-assemble into amyloid fibrils. These results prompt further studies to investigate the possible role of A{beta} condensates in the aggregation of this peptide in Alzheimers disease.

biophysics↗

bioRxiv2023

Ganglioside Lipids Inhibit the Aggregation of the Alzheimer's Related Peptide Amyloid-β

The aggregation of the amyloid-{beta} (A{beta}) peptides (A{beta}42/A{beta}40) into toxic amyloid fibrils and plaques is one of the molecular hallmarks in dementia and Alzheimers disease (AD). While the molecular mechanisms behind this aggregation process are not fully known, it has been shown that some biomolecules can accelerate this process while others can inhibit amyloid formation. Lipids, which are ubiquitously found in cell membranes, play a pivotal role in protein aggregation. Here, we investigate how ganglioside lipids, which are abundant in the brain and in neurons, can influence the aggregation kinetics of both A{beta}42 and A{beta}40. We find that ganglioside lipids can drastically inhibit the aggregation of A{beta}42, while in the case of the smaller peptide (A{beta}40), gangliosides can completely inhibit the aggregation process. Moreover, through kinetic analysis we show that the primary nucleation rate is greatly affected by the addition of gangliosides, and that the presence of these lipids can inhibit the primary nucleation rate of A{beta}42 by 3 orders of magnitude. By means of viability assays of neuroblastoma cells (SH-SY5Y), we further demonstrate that amyloid fibrils formed in the absence of gangliosides are more toxic to these cells than amyloid fibrils formed in the presence of gangliosides, elucidating the inhibitory and potentially protective role that these lipids can play. Additionally, we show that monomeric A{beta}40/A{beta}42 form complexes with gangliosides, but not with other lipids such as POPS, suggesting that formation of ganglioside-A{beta} complexes can act as a potential pathway towards inhibiting amyloid-{beta} aggregation. Taken together, our results provide a quantitative description of how lipid molecules such as gangliosides can inhibit the aggregation of A{beta} and shed light on the key factors that control these processes, especially in view of the fact that declining levels of gangliosides in neurons have been associated with ageing.

neuroscience↗

bioRxiv2022

Selenium Silk Nanostructured Films for Antifungal and Antibacterial Treatments

The rapid emergence of drug-resistant bacteria and fungi poses a threat for healthcare worldwide. The development of novel effective small molecule therapeutic strategies in this space has remained challenging. An orthogonal approach, therefore, is to explore biomaterials with physical modes of action which have the potential to generate antimicrobial activity and in some cases even prevent antimicrobial resistance. Here, to this effect, we describe an approach for forming silk-based films that contain embedded selenium nanoparticles. We show that these materials exhibit both antibacterial and antifungal properties while crucially also remaining highly biocompatible and non-cytotoxic towards mammalian cells. By incorporating the nanoparticles into silk films, the protein scaffold acts in a two-fold manner; it protects the mammalian cells from the cytotoxic effects of the bare nanoparticles, while also providing a template for bacterial and fungal eradication. A range of hybrid inorganic/organic films were produced and an optimum concentration was found, which allowed for both high bacterial and fungal death while also exhibiting low mammalian cell cytotoxicity. Such films can thus pave the way for next generation antimicrobial materials for applications such as wound healing and as agents against topical infections, with the added benefit that bacteria and fungi are unlikely to develop antimicrobial resistance to these hybrid materials.

microbiology↗

bioRxiv2022

Enhanced surface nano-analytics of transient biomolecular processes

The study of the physical and chemical properties of biomolecules enables the characterisation of fundamental molecular processes and mechanisms in health and disease. Bulk and single-molecule analytical methods provide rich information on biomolecules, but often require high concentrations and sample preparation away from physiologically relevant conditions. Here, we present the development and application of a lab-on-a-chip approach which combines rapid sample preparation, mixing and deposition to integrate with a range of nano-analytical methods in chemistry and biology, providing enhanced sensitivity and single molecule resolution. We demonstrate that this method empowers multidimensional study of heterogenous biomolecular systems in physiological buffers and concentrations over multiple length scales by nanoscopy and vibrational spectroscopy. We illustrate the capabilities of this platform by capturing and analysing the structural conformations of transient oligomeric species formed at the early stages of the self-assembly of -synuclein, which are associated with the onset of Parkinsons disease. TEASERMaintaining the heterogeneity and structural integrity of monomers and oligomers enables their quantitative study.

biophysics↗

bioRxiv2021

2-photon-fabricated nano-fluidic traps for extended detection of single macromolecules and colloids in solution

The analysis of nanoscopic species, such as proteins and colloidal assemblies, at the single-molecule level has become vital in many areas of fundamental and applied research. Approaches to increase the detection timescales for single molecules in solution without immobilising them onto a substrate surface and applying external fields are much sought after. Here we present an easy-to-implement and versatile nanofluidics-based approach that enables increased observational-timescale analysis of single biomacromolecules and nanoscale colloids in solution. We use two-photon-based hybrid lithography in conjunction with soft lithography to fabricate nanofluidic devices with nano-trapping geometries down to 100 nm in height. We provide a rigorous description and characterisation of the fabrication route that enables the writing of nanoscopic 3D structures directly in photoresist and allows for the integration of nano-trapping and nano-channel geometries within micro-channel devices. Using confocal fluorescence burst detection, we validated the functionality of particle confinement in our nano-trap geometries through measurement of particle residence times. All species under study, including nanoscale colloids, -synuclein oligomers, and double-stranded DNA, showed a three to five-fold increase in average residence time in the detection volume of nano-traps, due to the additional local steric confinement, in comparison to free space diffusion in a nearby micro-channel. Our approach thus opens-up the possibility for single-molecule studies at prolonged observational timescales to analyse and detect nanoparticles and protein assemblies in solution without the need for surface immobilisation.

biophysics↗

bioRxiv2020

Reentrant liquid condensate phase of proteins is stabilized by hydrophobic and non-ionic interactions

Many cellular proteins demix spontaneously from solution to form liquid condensates. These phase-separated systems have wide-ranging roles in health and disease. Elucidating the molecular driving forces underlying liquid-liquid phase separation (LLPS) is therefore a key objective for understanding biological function and malfunction. Here we show that proteins implicated in cellular LLPS, including FUS, TDP-43, Brd4, Sox2, and Annexin A11, which form condensates at low salt concentrations, can reenter a phase-separated regime at high salt concentrations. By bringing together experiments and simulations, we demonstrate that phase separation in the high-salt regime is driven by hydrophobic and non-ionic interactions, and is mechanistically distinct from the low-salt regime, where condensates are additionally stabilized by electrostatic forces. Our work thus provides a new view on the cooperation of hydrophobicity and non-ionic interactions as non-specific driving forces for the condensation process, with important implications for aberrant function, druggability, and material properties of biomolecular condensates.

biophysics↗