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March 1998 - present
Publications
Publications (178)
The neurotransmitter serotonin is involved in physiological processes such as appetite, sleep, and mood and diseases such as anxiety and depression. Traditionally, the effects of serotonin were thought to be initiated by binding to its target transmembrane receptors. It is also known that serotonin can bind directly to the membrane with high affini...
Designing functional molecules which can recognize and modify the activity of a specific protein is a frequently encountered challenge in biology and pharmaceutical chemistry, and requires major effort for each specific protein target. Here we demonstrate that “self-peptides”, parts of folded proteins which by their nature are recognizable by the r...
Adult hippocampal neurogenesis is a lifelong process that involves the integration of newborn neurons into the hippocampal network, and plays a role in cognitive function and the modulation of mood-related behavior. Here, we sought to address the impact of chemogenetic activation of adult hippocampal progenitors on distinct stages of progenitor dev...
The sheet‐like lipid bilayer is the fundamental structural component of all cell membranes. Its building blocks are phospholipids and cholesterol. Their amphiphilic structure spontaneously leads to the formation of a bilayer in aqueous environment. Lipids are not just structural elements. Individual lipid species, the lipid membrane structure, and...
The interaction of disease-causing amyloid oligomers with lipid membranes is implicated in their toxicity. However, understanding the membrane interaction of different oligomers, and each constituent monomer in a given oligomer, has remained a challenge. Here we employed a recently developed single-molecule technique, called QSLIP, which can simult...
The primary event in chemical neurotransmission involves the fusion of a membrane-limited vesicle at the plasma membrane and the subsequent release of its chemical neurotransmitter cargo. The cargo itself is not known to have any effect on the fusion event. However, amphiphilic monoamine neurotransmitters (e.g. serotonin and dopamine) are known to...
Liquid−liquid phase separation (LLPS) plays a crucial role in cellular organization, primarily driven by intrinsically disordered proteins (IDPs) leading to the formation of biomolecular condensates. A folded protein SUMO that post-translationally
modifies cellular proteins has recently emerged as a regulator of LLPS. Given its compact structure an...
It is poorly understood why ApoE variants are major genetic risk factors in Alzheimer’s disease (AD), which is associated with the aggregation of amyloid beta (Aβ). Here we directly image specific changes in small Aβ oligomers in rat brain cells that correlate with the cellular ApoE content. An inhibitor of Aβ-ApoE interaction suppresses this chang...
Adult hippocampal neurogenesis is a lifelong process that involves the integration of newborn neurons into the hippocampal network, and plays a role in cognitive function and the modulation of mood-related behavior. Here, we sought to address the impact of chemogenetic activation of adult hippocampal progenitors on distinct stages of progenitor dev...
Many studies in recent years have investigated the phenomenon of liquid-liquid phase separation (LLPS) in proteins. LLPS is reported in intrinsically disordered proteins (IDPs) or in proteins with intrinsically disordered regions (IDRs) lacking a well-defined three-dimensional structure. However, the occurrence of LLPS in folded proteins, that lack...
Nature confines hundreds of millimolar of amphiphilic neurotransmitters, such as serotonin, in synaptic vesicles. This appears to be a puzzle, as the mechanical properties of lipid bilayer membranes of individual major polar lipid constituents of synaptic vesicles [phosphatidylcholine (PC), phosphatidylethanolamine (PE), and phosphatidylserine (PS)...
Allelic variants of ApoE are the most significant genetic risk factors for late-onset Alzheimer’s disease, which is likely caused by the aggregation of Amyloid-β (Aβ) in the brain. However, understanding how the interaction between these two species lead to toxicity has remained a major challenge. APoE seems to affect the aggregation of Aβ in vitro...
Small lipid vesicles (with diameter ≤100 nm) with their highly curved membranes comprise a special class of biological lipid bilayers. The mechanical properties of such membranes are critical for their function, e.g. exocytosis. Cholesterol is a near-universal regulator of membrane properties in animal cells. Yet measurements of the effect of chole...
Small oligomers are the initial intermediates in the pathway to amyloid fibril formation. They have a distinct identity from the monomers as well as from the protofibrils and the fibrils, both in their structure and in their properties. In many cases, they play a crucial biological role. However, due to their transient nature, they are difficult to...
‘Membrane order’ is a term commonly used to describe the elastic and mechanical properties of the lipid bilayer, though its exact meaning is somewhat context- and method-dependent. These mechanical properties of the membrane control many cellular functions and are measured using various biophysical techniques. Here we ask if the results obtained fr...
Knowledge of the excitation profile in a confocal or multiphoton microscope can improve the image resolution, e.g. by using deconvolution, pixel reassignment or adaptive optics strategies. Here we demonstrate a method by which the scanning beam can be used to place a stationary, virtual ‘guide star’ at any chosen location in the sample, during imag...
Serotonin is an endogenous neurotransmitter involved in both physiological and pathophysiological processes. Traditionally, serotonin acts as a ligand for G protein-coupled receptors (GPCRs) leading to subsequent cell signaling. However, serotonin can also bind to lipid membranes with high affinity and modulate the phase behavior in 1-palmitoyl-2-o...
Small oligomers are widely accepted to be the major toxic agents in amyloid diseases. However, it is difficult to decipher their structure or conformation. Small oligomers of different sizes are in dynamic equilibrium with each other, and are therefore not amenable to separation techniques. Single molecule approaches can study individual members of...
Bacterial amyloids decorate the cell surface of many bacteria by forming functional amyloid fibers. These amyloids have structural and biochemical similarities with many disease-related amyloids in eukaryotes. Amyloid aggregation starts at the individual monomer level, and the end product is the amyloid fibril. The process of amyloid aggregation in...
Amyloid β (Aβ) is a peptide known to form amyloid fibrils in the brain of patients suffering from Alzheimer’s disease. A complete mechanistic understanding how Aβ peptides form neurotoxic assemblies and how they kill neurons has not yet been achieved. Previous analysis of various Aβ40 mutants could reveal the significant importance of the hydrophob...
An important measure of the conformation of protein molecules is the degree of surface exposure of its specific segments. However, this is hard to measure at the level of individual molecules. Here, we combine single molecule photobleaching (smPB, which resolves individual photobleaching steps of single molecules) and fluorescence quenching techniq...
The entry of the SARS-CoV2 virus in human cells is mediated by the binding of its surface spike protein to the human Angiotensin-Converting Enzyme 2 (ACE2) receptor. A 23 residues long helical segment (SBP1) at the binding interface of human ACE2 interacts with viral spike protein and therefore, has generated considerable interest as a recognition...
Serotonin is an important signaling molecule with a high lipid membrane affinity. Membrane binding of serotonin modulates membrane properties and potentially cellular function as well, providing an activity pathway completely independent of serotonin receptors. The results presented in this study suggest a mode of serotonin–membrane interaction tha...
Serotonin, an important signaling molecule in humans, has an unexpectedly high lipid membrane affinity. The significance of this finding has evoked considerable speculation. Here we show that membrane binding by serotonin can directly modulate membrane properties and cellular function, providing an activity pathway completely independent of seroton...
Unsaturated and saturated phospholipids tend to laterally segregate, especially in the presence of cholesterol. Small molecules such as neurotransmitters, toxins, drugs etc. possibly modulate this lateral segregation. The small aromatic neurotransmitter serotonin (5-HT) has been found to bind to membranes. We studied the lipid structure and packing...
Neuronal plaques of amyloid β (Aβ) peptides of varying length carrying different posttranslational modifications represent a molecular hallmark of Alzheimer’s disease. It is believed that transient oligomeric Aβ assemblies associating in early fibrillation events represent particularly cytotoxic peptide aggregates. Also, N-terminally truncated (in...
Serotonin is a neurotransmitter as well as a somatic signaling molecule, and the serotonergic system is a major target for psychotropic drugs. Serotonin, together with a few related neurotransmitters, has recently been found to exhibit an unexpectedly high lipid membrane affinity ¹⁻³ . It has been conjectured that extrasynaptic serotonin can diffus...
Oligomers are the key suspects in protein aggregation-linked diseases, such as Alzheimer’s and Type II diabetes, and most likely exert their toxicity by interacting with lipid membranes. However, the “which oligomer” question remains an obstacle in understanding the disease mechanism, as the exact identity of the toxic oligomer(s) is not yet known....
Protein folding can go wrong in vivo and in vitro, with significant consequences for the living cell and the pharmaceutical industry, respectively. Here we propose a general design principle for constructing small peptide-based protein-specific folding modifiers. We construct a xenonucleus, which is a pre-folded peptide that resembles the folding n...
An amyloid aggregate evolves through a series of intermediates which have different secondary structures and intra/intermolecular contacts. The structural parameters of these intermediates are important determinants of their toxicity. For example, the early oligomeric species of the amyloid β (Aβ) peptide have been implicated as the most cytotoxic...
Single molecule photobleaching is a powerful technique to measure the number of fluorescent units in subresolution molecular complexes, such as in toxic protein oligomers associated with amyloid diseases. However, photobleaching can occur before the sample is appropriately placed and focused. Such "prebleaching" can introduce a strong systematic bi...
Designer receptors exclusively activated by designer drugs (DREADD)-based chemogenetic tools are extensively used to manipulate neuronal activity in a cell type-specific manner. Whole-cell patch-clamp recordings indicate membrane depolarization, coupled with increased neuronal firing rate, following administration of the DREADD ligand, clozapine-N-...
Designer Receptors Exclusively Activated by Designer Drugs (DREADD)-based chemogenetic tools are extensively used to manipulate neuronal activity in a cell-type specific manner. Whole-cell patch-clamp recordings indicate membrane depolarization, coupled with increased neuronal firing rate, following administration of the DREADD ligand, Clozapine-N-...
While the roles of intrinsically disordered protein domains in driving many interactions are increasingly well‐appreciated, the mechanism of toxicity of disease‐causing disordered proteins remains poorly understood. A prime example is Alzheimer's disease (AD) associated amyloid beta (Aβ). Aβ oligomers are highly toxic partially structured peptide a...
While the roles of intrinsically disordered protein domains in driving many interactions are increasingly well-appreciated, the mechanism of toxicity of disease-causing disordered proteins remains poorly understood. A prime example is Alzheimer’s disease (AD) associated amyloid beta (Aβ). Aβ oligomers are highly toxic partially structured peptide a...
It is about designing a 'decoy folding nucleus' that can modulate protein folding kinetics.
Visualizing small biomolecules in living cells remains a difficult challenge. Neurotransmitters provide one of the most frustrating examples of this difficulty, as our understanding of signaling in the brain critically depends on our ability to follow the neurotransmitter traffic. Last two decades have seen considerable progress in probing some of...
Surface enhanced Raman spectroscopy (SERS) has great potential for investigating the secondary structure of biomolecules. While enhancements of many orders of magnitude are possible in SERS, the excitation powers required for adequate signal to noise may still be damaging for sensitive protein molecules. Here, we describe the design and constructio...
We demonstrate a technique which can change the pH inside an NMR tube in steps of ≤0.01 pH unit, during a solution-state NMR experiment. We use a photo-acid co-solute with the actual specimen, and photo-excite the solution by direct fibre-mediated light delivery to the NMR tube. The photo-excitation and NMR measurements can be performed simultaneou...
Single-molecule photobleaching (smPB) technique is a powerful tool for characterizing molecular assemblies. It can provide a direct measure of the number of monomers constituting a given oligomeric particle and generate the oligomer size distribution in a specimen. A major current application of this technique is in understanding protein aggregatio...
Short range plasmonic fields around a nanoparticle can modulate fluorescence or Raman processes. In lipid encased nanoparticles, this can potentially measure the relative depths of different parts of a membrane...
We investigated the influence of the chemical structure of the phenylalanine side chain in position 19 of the 40 residue amyloid β peptide. Side chain modifications in this position yielded...
Islet amyloid polypeptide (IAPP) is a 37 residue intrinsically disordered protein whose aggregation is associated with Type II diabetes. Like most amyloids, it appears that the intermediate aggregates ("oligomers") of IAPP are more toxic than the mature fibrils, and interaction with the cell membrane is likely to be an integral component of the tox...
Membrane interaction appears to be a key step in the toxic pathway of many amyloidogenic proteins, such as Amyloidβ (Aβ 40/42 , associated with Alzheimer's disease) and hIAPP (associated with Type II diabetes). While the oligomers are generally considered to be more toxic than either the monomers or the fibrils, the exact identity of the toxic olig...
The formation of the hydrophobic contact between phenylalanine 19 (F19) and leucine 34 (L34) of amyloid β (1-40) (Aβ(1-40)) is known to be an important step in the fibrillation of Aβ(1-40) peptides. Mutations of this putatively early molecular contact were shown to strongly influence the toxicity of Aβ(1-40) (Das et al., ACS Chem. Neurosci. 6 (2015...
Monoamine neurotransmission is key to neuromodulation, but imaging monoamines in live neurons has remained a challenge. Here we show that externally added ortho-phthalaldehyde (OPA) can permeate live cells and form bright fluorogenic adducts with intracellular monoamines (e.g. setrotonin, dopamine and nor-epinephrine) and with L-DOPA, which can be...
Ratiometric imaging can quantitatively measure changes in cellular analyte concentrations using specially designed fluorescent labels. We describe a label-free ratiometric imaging technique for direct detection of changes in intravesicular serotonin concentration in live cells. At higher concentrations, serotonin forms transient oligomers whose ult...
The structural underpinnings for the higher toxicity of the oligomeric intermediates of amyloidogenic peptides, compared to the mature fibrils, remain unknown at present. The transient nature and heterogeneity of the oligomers make it difficult to follow their structure. Here, using vibrational and solid-state nuclear magnetic resonance spectroscop...
There are three specific regions in the Amyloid beta (Aβ) peptide sequence where variations cause enhanced toxicity in Alzheimer’s disease: the N-terminus, the central salt bridge, and the C-terminus. Here, we investigate if there is a close conformational connection between these three regions, which may suggest a concerted mechanism of toxicity....
Small oligomers are the major toxic species in many amyloid related diseases, but they are difficult to characterize and target. Here we construct tetra-peptides FXFX (X= F/K), designed to exploit cation-π, π-π and hydrophobic interactions to disrupt the critical F19-L34 contact recently found in Aβ40 oligomers. FRFR accelerates Aβ40 aggregation, a...
The Inside Back Cover summarizes the strategy applied to study the influence of local physical forces on the fibrillation kinetics, structure, and dynamics of amyloid β (1–40) fibrils. Rationally designed mutations were introduced to modify the hydrophobic contact between residues 19 and 34. More information can be found in the Full Paper by J. Adl...
We have generated a small library of rationally designed amyloid β (Aβ(1-40)) peptide variants and studied the morphology of the fibrils formed by these peptide variants. In these molecules, the structurally important hydrophobic contact between phenylalanine 19 (F19) and leucine 34 (L34) was systematically mutated to introduce defined physical for...
Toxicity of Aβ aggregates, suspected to be the key to Alzheimer's disease, is strongly size dependent. Toxicity is expected to be related to structure, which possibly evolves with size. However, structural studies have suggested that the structured parts of the peptide remain rather similar between the oligomers and the mature fibrils, and are domi...
: Aggregation of human amylin, a 37 amino acid residue neuropeptide of pancreatic origin, into amyloid aggregates is implicated in the etiology of diabetes mellitus type II. Despite its clinical significance, details of progression of nontoxic amylin monomers into cytotoxic oligomers are not very clear. Studies based on Amyloid-β (Aβ), a peptide wi...
Protein aggregation poses a fundamental problem in biophysics, whose solutions have enormous potential for societal benefits. Many devastating and incurable diseases, such as Alzheimer’’s, Parkinson’’s and Type II diabetes, are strongly linked to the misfolding and aggregation of specific proteins. The links between misfolding, aggregation and toxi...
Aβ self-assembles into parallel cross-β fibrillar aggregates, which is associated with Alzheimer's disease pathology. A central
hairpin turn around residues 23–29 is a defining characteristic of Aβ in its aggregated state. Major biophysical properties
of Aβ, including this turn, remain unaltered in the central fragment Aβ18–35. Here, we synthesize...
Identifying the structures of membrane bound proteins is critical to understanding their function in healthy and diseased states. We introduce a surface enhanced Raman spectroscopy technique which can determine the conformation of membrane-bound proteins, at low micromolar concentrations, and also in the presence of a substantial membrane-free frac...
Certain neurodegenerative diseases are thought to be initiated by the aggregation of amyloidogenic proteins. However, the mechanism underlying toxicity remains obscure. Most of the suggested mechanisms are generic in nature and do not directly explain the neuron-type specific lesions observed in many of these diseases. Some recent reports suggest t...
Small hydrophobic oligomers of aggregation-prone proteins are thought to be generically toxic. Here we examine this view by perturbing an early folding contact between Phe19 and Leu34 formed during the aggregation of Alzheimer's amyloid-β (Aβ40) peptide. We find that even conservative single mutations altering this interaction can abolish Aβ40 toxi...
Small oligomers of Amyloid-β (Aβ), rather than the mature fibrils, are suspected to initiate Alzheimer's disease (AD). The structures of these oligomers in the solution, and in the membrane phase, are expected to provide significant cues for addressing their toxicity. However, it is difficult to determine the structure of these peptides as they are...
Small oligomers of Amyloid beta (Aβ) are suspected to be the key to Alzheimer disease (AD). However, identifying these toxic species in the background of other similar but non-toxic Aβ aggregates has remained a challenge. Recent studies indicate that Aβ undergoes a global structural transition at an early step of aggregation. This transition is mar...
Small oligomers of the amyloid β (Aβ) peptide, rather than the monomers or the fibrils, are suspected to initiate Alzheimer′s disease (AD). However, their low concentration and transient nature under physiological conditions have made structural investigations difficult. A method for addressing such problems has been developed by combining rapid fl...
Small oligomers of the amyloid β (Aβ) peptide, rather than the monomers or the fibrils, are suspected to initiate Alzheimer′s disease (AD). However, their low concentration and transient nature under physiological conditions have made structural investigations difficult. A method for addressing such problems has been developed by combining rapid fl...
Dopaminergic neurotransmission has been investigated extensively, yet direct optical probing of dopamine has not been possible in live cells. Here we image intracellular dopamine with sub-micron three-dimensional resolution by harnessing its intrinsic mid-ultraviolet (UV) autofluorescence. Two-photon excitation with visible light (540 nm) in conjun...
Amyloid β (Aβ) fibrillar deposits in the brain are a hallmark of Alzheimer disease (AD). Curcumin, a common ingredient of
Asian spices, is known to disrupt Aβ fibril formation and to reduce AD pathology in mouse models. Understanding the structural
changes induced by curcumin can potentially lead to AD pharmaceutical agents with inherent bio-compat...
Alzheimer's Amyloid-β (Aβ) peptide changes its conformation as it aggregates (1) and acquires toxicity, ultimately causing Alzheimer's disease. Since it is difficult to decouple conformational changes from aggregation, any cause-effect relationship between them remains poorly explored. Here we attach Alzheimer's Amyloid-β40 monomers to silver nanop...
Decoupling conformational changes from aggregation will help us understand amyloids better. Here we attach Alzheimer's amyloid-beta(1-40) monomers to silver nanoparticles, preventing their aggregation, and study their conformation under aggregation-favoring conditions using SERS. Surprisingly, the alpha-helical character of the peptide remains unch...
Decoupling conformational changes from aggregation will help us understand amyloids better. Here we attach Alzheimer’s Amyloid-β1-40 monomers to silver nanoparticles, preventing their aggregation, and study their conformation under aggregation-favoring conditions using SERS. Surprisingly, the α-helical character of the peptide remains unchanged bet...
Small amyloid-β (Aβ) oligomers have much higher membrane affinity compared to the monomers, but the structural origin of this functional change is not understood. We show that as monomers assemble into small n-mers (n < 10), Aβ acquires a tertiary fold that is consistent with the mature fibrils. This is an early and defining transition for the aggr...
