Coronavirus disease 2019 (COVID-19) pandemic caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has become a global crisis. There is no therapeutic treatment specific for COVID-19. It is highly desirable to identify potential antiviral agents against SARS-CoV-2 from existing drugs available for other diseases and, thus, repurpose them for treatment of COVID-19. In general, a drug

Highly sensitive detection of proteins is of central importance to biomolecular analysis and diagnostics. Conventional protein sensing assays, such as ELISAs, remain reliant on surface-immobilization of target molecules and multi-step washing protocols for the removal of unbound affinity reagents. These features constrain parameter space in assay design, resulting in fundamental limitations due to

The quantification of transport processes of different substances in the brain's parenchyma is important in the context of understanding brain functioning. Most of the currently used methods for assessment of the effective diffusion coefficient rely on the point-source paradigm. We propose a method for the quantitative characterization of the diffusion process in the brain's parenchyma using a set

COVID-19, caused by SARS-CoV-2, lacks effective therapeutics. Additionally, no antiviral drugs or vaccines were developed against the closely related coronavirus, SARS-CoV-1 or MERS-CoV, despite previous zoonotic outbreaks. To identify starting points for such therapeutics, we performed a large-scale screen of electrophile and non-covalent fragments through a combined mass spectrometry and X-ray approach

Living systems at all scales aggregate in large numbers for a variety of functions including mating, predation, and survival. The majority of such systems consist of unconnected individuals that collectively flock, school or swarm. However some aggregations involve physically entangled individuals, which can confer emergent mechanofunctional material properties to the collective. Here we study in laboratory

Since its initial development in 1976, fluorescence recovery after photobleaching (FRAP) has been one of the most popular tools for studying diffusion and protein dynamics in living cells. Its popularity is derived from the widespread availability of confocal microscopes and the relative ease of the experiment and analysis. FRAP, however, is limited in its ability to resolve spatial heterogeneity.

Coronaviruses may produce severe acute respiratory syndrome (SARS). Indeed, a new SARS-type virus, SARS-CoV-2, is responsible of a global pandemic in 2020 with unprecedented sanitary and economic consequences for most countries. In the present contribution we study, by using all-atom equilibrium and enhanced sampling molecular dynamics simulations, the interaction between the SARS Unique Domain and

Mechanistic understanding of DNA recombination in the Cre-loxP system has largely been guided by crystallographic structures of tetrameric synaptic complexes. Those studies have suggested a role for protein conformational dynamics that has not been well characterized at the atomic level. We used solution NMR to discover the link between intrinsic flexibility and enzyme function in Cre recombinase.

An enhanced conformational sampling, a genetic-algorithm-based multidimensional virtual-system coupled molecular dynamics (GA-based mD-VcMD), was applied to a peptide, phosphorylated Myeloid leukemia factor 1 (pMLF1), binding to a receptor protein, 14-3-3ε, in an explicit solvent. This method provides an equilibrated ensemble at room temperature, from which a density map of pMLF1 around 14-3-3ε can

We study the structural ensembles of human chromosomes across different cell types. Using computer simulations, we generate cell-specific 3D chromosomal structures and compare them to recently published chromatin structures obtained through super-resolution microscopy. We demonstrate using a combination of machine learning and polymer physics simulations that epigenetic information can be used to predict

In recent years, the observed antibody sequence space has grown exponentially due to advances in high-throughput sequencing of immune receptors. The rise in sequences has not been mirrored by a rise in structures, as experimental structure determination techniques have remained low-throughput. Computational modeling, however, has the potential to close the sequence–structure gap. To achieve this goal

In this study we examined the influence of a transmembrane voltage on the hydrophobic gating of nanopores using molecular dynamics simulations. We observed electric field induced wetting of a hydrophobic gate in a biologically inspired model nanopore based on the 5-HT3 receptor in its closed state, with a field of at least ∼100 mV nm−1 was required to hydrate the pore. We also found an unequal distribution

SYBR Gold is a commonly used and particularly bright fluorescent DNA stain, however, its binding mode to DNA remains controversial. Here, we quantitate SYBR Gold binding to DNA using two complementary approaches. We use mechanical micromanipulation with magnetic tweezers (MT) to determine the effects of SYBR Gold binding on DNA length, twist, and mechanical properties. The MT assay reveals systematic

Extracellular vesicles (EVs) represent a potent intercellular communication system. Within a lipid bilayer such small vesicles transport biomolecules between cells and throughout the body, strongly influencing the fate of recipient cells. Due to their specific biological functions they have been proposed as biomarkers for various diseases and as optimal candidates for therapeutic applications. Despite

The scission of lipid membranes is a common biological process, often mediated by ESCRT complexes in concert with VPS4 assembling around the separation point. The functions of the ESCRT-I and ESCRT-III complexes are well established in certain of these cellular processes; however, the role of ESCRT- II remains contentious. Here, we devised a SNAP-tag fluorescent labelling strategy to understand the

The molecular chaperone 90-kDa heat-shock protein (Hsp90) assists the late-stage folding and activation of diverse types of protein substrates (called clients), including many kinases. Previous studies have established that the Hsp90 homodimer undergoes an ATP-driven cycle through open and closed conformations. Here I propose a model of client activation by Hsp90, which predicts that this cycle enables

Glutamate is the most abundant excitatory neurotransmitter in the central nervous system, therefore its precise control is vital for maintaining normal brain function and preventing excitotoxicity1. Removal of extracellular glutamate is achieved by plasma membrane-bound transporters, which couple glutamate transport to sodium, potassium and pH gradients using an elevator mechanism2-5. Glutamate transporters

During clathrin-mediated endocytosis, a patch of flat plasma membrane is deformed into a vesicle. In walled cells, such as plants and fungi, the turgor pressure is high and pushes the membrane against the cell wall, thus hindering membrane internalization. In this paper, we study how a patch of membrane is deformed against turgor pressure by force and by curvature-generating proteins. We show that

Here, we used for the first time Hydrogen-Deuterium eXchange coupled to Mass Spectrometry (HDX-MS) to investigate conformational differences between the human standard 20S (std20S) and immuno 20S (i20s) proteasomes alone or in complex with PA28αβ or PA28γ activators. Their solvent accessibility was analyzed through a dedicated bioinformatic pipeline including stringent statistical analysis and 3D visualization

Cell division of Staphylococcus adopts a 'popping' mechanism that mediates extremely rapid separation of the septum. Elucidating the structure of the septum is crucial for understanding this exceptional bacterial cell division mechanism. Here, the septum structure of Staphylococcus warneri is extensively characterized using high-speed time-lapse confocal microscopy, atomic force microscopy, and electron

Plexins receive guidance cues from semaphorin ligands and transmit their signal through the plasma membrane. This family of proteins is unique amongst single-pass transmembrane receptors as their intracellular regions interact directly with several small GTPases, which regulate cytoskeletal dynamics and cell adhesion. Here, we characterize the GTPase Activating Protein (GAP) function of Plexin-B1 and

Thylakoid membranes in chloroplasts contain photosynthetic protein complexes that convert light energy into chemical energy. Photosynthetic protein complexes are considered to undergo structural reorganization to maintain the efficiency of photochemical reactions. A detailed description of the mobility of photosynthetic complexes in real-time is necessary to understand how macromolecular organization

Mechanical cell competition is important during tissue development, cancer invasion, and tissue ageing. Heterogeneity plays a key role in practical applications since cancer cells can have different cell stiffness and different proliferation rates than normal cells. To study this phenomenon, we propose a one-dimensional mechanical model of heterogeneous epithelial tissue dynamics that includes cell-length-dependent

Key enzymatic processes in biology use the nonequilibrium error correction mechanism called kinetic proofreading to enhance their specificity. Kinetic proofreading typically requires several dedicated structural features in the enzyme, such as a nucleotide hydrolysis site and multiple enzyme-substrate conformations that delay product formation. Such requirements limit the applicability and the adaptability

Molecular segregation and biopolymer manipulation require the action of molecular motors to do work by applying directional forces to macromolecules. The additional strand conserved E (ASCE) ring motors are an ancient family of molecular motors responsible for diverse tasks ranging from biological polymer manipulation (e.g. protein degradation and chromosome segregation) to establishing and maintaining

The contraction of contractile rings (CRs) depends on interaction between actin filaments and myosin II filaments. The rate of contraction in the fission yeast Schizosaccharomyces pombe is less than 1/120 of the velocity of acto-myosin II movement in vitro, but the mechanism of inhibition has not been described. Here, we found that the calponin-homology actin binding domain of fission yeast IQGAP Rng2

We demonstrate a combined experimental and computational approach for the quantitative characterization of lateral interactions between membrane-associated proteins. In particular, weak, lateral (cis) interactions between E-cadherin extracellular domains tethered to supported lipid bilayers, were studied using a combination of dynamic single-molecule Förster Resonance Energy Transfer (FRET) and kinetic

N-glycosylation is one of the most abundant and diverse post-translational modifications of proteins, implicated in protein folding and structural stability, and mediating interactions with receptors and with the environment. All N-glycans share a common core from which linear or branched arms stem from, with functionalization specific to different species and to the cells' health and disease state

Human carbonic anhydrase 1 (CA1) has been suggested as a biomarker for identification of several diseases including cancers, pancreatitis, diabetes, and Sjogren's syndrome. However, the lack of a rapid, cheap, accurate, and easy-to-use quantifcation technique has prevented widespread utilization of CA1 for practical clinical applications. To this end, we present a label-free electronic biosensor for

E-cadherin adhesions are essential for cell-to-cell cohesion and mechanical coupling between epithelial cells and reside in a micro-environment that comprises the adjoining epithelial cells. While E-cadherin has been shown to be a mechanosensor, it is unknown if E-cadherin adhesions can differentially sense stiffness within the range of that of epithelial cells. A survey of literature shows that epithelial

During the multi-color fluorescence imaging, a fluorophore can be detected in the non-corresponding channel due to the broad spectra of the fluorophore and filters, resulting in the artefact called the crosstalk (crossover or bleed-through). Unfortunately, the fluorescence crosstalk is ubiquitous for the conventional fluorescence microscopy. Significant crosstalk can distort the image and lead to incorrect

Dense surface glycosylation on the HIV-1 envelope (Env) protein acts as a shield from the adaptive immune system. However, the molecular complexity and flexibility of glycans make experimental studies a challenge. Here we have integrated high-throughput atomistic modeling of fully glycosylated HIV-1 Env with graph theory to capture immunologically important features of the shield topology. This is

Most foldable protein sequences adopt only a single native fold. Recent protein design studies have, however, created protein sequences which fold into different structures apon changes of environment, or single point mutation, the best characterized example being the switch between the folds of the GA and GB binding domains of streptococcal protein G. To obtain further insight into the design of sequences

Toehold-mediated strand displacement (TMSD) is a nucleic acid-based reaction wherein an invader strand (I) replaces an incumbent strand (N) in a duplex with a target strand (T). TMSD is driven by toeholds, overhanging single-stranded domains in T recognised by I. Although TMSD is responsible for the outstanding potential of dynamic DNA nanotechnology, TMSD cannot implement templating, the central mechanism

Biomolecules do not fold into a single 3D structure but rather form dynamic ensembles of many inter-converting conformations. Knowledge of dynamic ensembles is key for understanding how biomolecules fold and function, and for rationally manipulating their activities in drug discovery and synthetic biology. However, solving dynamic ensembles of biomolecules at atomic resolution is a major challenge

Ring ATPases that translocate disordered polymers possess lock-washer architectures that they impose on their substrates during transport via a hand-over-hand mechanism. Here, we investigate the operation of ring motors that transport substrates possessing a preexisting helical structure, such as the bacteriophage φ29 dsDNA packaging motor. During each cycle, this pentameric motor tracks one helix

The three-dimensional positions of atoms in protein molecules define their structure and provide mechanistic insights into the roles they perform in complex biological processes. The more precisely atomic coordinates are determined, the more chemical information can be derived and the more knowledge about protein function may be inferred. With breakthroughs in electron detection and image processing

Plant cell division involves de novo formation of a cell plate that partitions the cytoplasm of the dividing cell. Cell plate formation is directed by orchestrated delivery of cytokinetic vesicles via the phragmoplast, vesicle fusion, and membrane maturation to the nascent cell wall by the timely deposition of polysaccharides such as callose, cellulose, and crosslinking glycans. In contrast to the

The probabilities that two loci in chromosomes that are separated by a certain genome length can be inferred using chromosome conformation capture method and related Hi-C experiments. How to go from such maps to an ensemble of three-dimensional structures, which is an important step in understanding the way nature has solved the packaging of the hundreds of million base pair chromosomes in tight spaces

Podocytes are crucial cells of the glomerular filtration unit and playing a vital role at the interface of the blood-urine barrier. Podocyte slit-diaphragm is a modified tight junction that facilitates size and shape-dependent permselectivity. Several proteins including podocin, nephrin, CD2AP, and TRPC6 form a macromolecular assembly and constitute the slit-diaphragm. Podocin is an integral membrane

Membrane bending is a ubiquitous cellular process that is required for membrane traffic, cell motility, organelle biogenesis, and cell division. Proteins that bind to membranes using specific structural features, such as wedge-like amphipathic helices and crescent-shaped scaffolds, are thought to be the primary drivers of membrane bending. However, many membrane-binding proteins have substantial regions

Many immunoreceptors have cytoplasmic domains that are intrinsically disordered (i.e., have high configurational entropy), have multiple sites of post-translational modification (e.g., tyrosine phosphorylation), and participate in nonlinear signaling pathways (e.g., exhibiting switch-like behavior). Several hypotheses to explain the origin of these nonlinearities fall under the broad hypothesis that

Cyclooxygenases carry out the committed step in prostaglandin synthesis and are the target of NSAIDs, the most widely used class of drugs in alleviating pain, fever, and inflammation. While extensively studied, one aspect of their biology that has been neglected is their interaction with membrane lipids. Such lipid-protein interactions have been shown to be a driving force behind membrane protein function

Purpose: Intervertebral disc degeneration (IVDD), resulting in the depletion of hydrophilic glycosaminoglycans (GAGs) located in the nucleus pulposus (NP), can lead to debilitating neck and back pain. Magnetic Resonance Imaging (MRI) is a promising means of IVD assessment due to the sensitivity of MRI tissue relaxation properties to matrix composition. Furthermore, anomalous (i.e. non-monoexponential)

The SpCas9 endonuclease has become an important tool in gene-editing and basic science alike. Though easily programmed to target any sequence, SpCas9 also shows considerable activity over genomic off-targets. Many empirical facts regarding the targeting reaction have been established, but a comprehensive mechanistic description is still lacking - limiting fundamental understanding, our ability to predict

Riboswitches are regulatory ribonucleic acid (RNA) elements that act as ligand-dependent conformational switches. In the apo form, the aptamer domain, the region of a riboswitch that binds to its cognate ligand, is dynamic, thus requiring an ensemble-representation of its structure. Analysis of such ensembles can provide molecular insights into the sensing mechanism and capabilities of riboswitches

Cells interact mechanically with their surrounding by exerting forces and sensing forces or force-induced displacements. Traction force microscopy (TFM), purported to map cell-generated forces or stresses, represents an important tool that has powered the rapid advances in mechanobiology. However, to solve the ill-posted mathematical problem, its implementation has involved regularization and the associated

We measured viscoelasticity of single protein molecules using two types of Atomic Force Microscopes (AFM) which employ different detection schemes to measure the cantilever response. We used a commonly available deflection detection scheme in commercial AFMs which measures cantilever bending and a fibre-interferometer based home-built AFM which measures cantilever displacement. For both methods, the

Methods to directly inhibit gene expression using small molecules hold promise for the development of new therapeutics targeting proteins that have evaded previous attempts at drug discovery. Among these, small molecules including the drug-like compound PF- 06446846 (PF846) selectively inhibit the synthesis of specific proteins, by stalling translation elongation. These molecules also inhibit translation

Phenomenological relations such as Ohm's or Fourier's law have a venerable history in physics, but are still scarce in biology. This situation restrains predictive theory. Here, we build on bacterial "growth laws," which capture physiological feedback between translation and cell growth, to construct a minimal biophysical model for the combined action of ribosome-targeting antibiotics. Our model predicts

We introduce single molecule light field microscopy (SMLFM), a novel 3D single molecule localization technique that is capable of up to 20 nm isotropic precision across a 6 μm depth of field. SMLFM can be readily implemented by installing a refractive microlens array into the conjugate back focal plane of any widefield single molecule localization system. We demonstrate that 3D localization can be

Single particle cryo-EM is a powerful method to solve the three-dimensional structures of biological macromolecules. The technological development of electron microscopes, detectors, automated procedures in combination with user friendly image processing software and ever-increasing computational power have made cryo-EM a successful and largely expanding technology over the last decade. At resolutions

A widely used method to measure the bending rigidity of bilayer membranes is fluctuation spectroscopy, which analyses the thermally-driven membrane undulations of giant unilamellar vesicles recorded with either phase-contrast or confocal microscopy. Here, we analyze the fluctuations of the same vesicle using both techniques and obtain consistent values for the bending modulus. We discuss the factors

DNA strand displacement, where a single-stranded nucleic acid invades a DNA duplex, is pervasive in genomic processes and DNA engineering applications. The kinetics of strand displacement have been studied in bulk; however, the kinetics of the underlying strand exchange were obfuscated by a slow bimolecular association step. Here, we use a novel single-molecule Fluorescence Resonance Energy Transfer

Mammalian sirtuins (SIRT1-SIRT7) are a family of nicotinamide adenine dinucleotide (NAD+)-dependent protein deacylases that play critical roles in lifespan and age-related diseases. The physiological importance of sirtuins has stimulated intense interest in designing sirtuin-activating compounds. However, except for allosteric activators of SIRT1-catalyzed reactions that are limited to particular substrates

Protein disorder and aggregation play significant roles in the pathogenesis of numerous neurodegenerative diseases, such as Alzheimer's and Parkinson's disease. The end products of the aggregation process in these diseases are β-sheet rich amyloid fibrils. Though in most cases small, soluble oligomers formed during amyloid aggregation are the toxic species. A full understanding of the physicochemical

Clustering of proteins is crucial for many cellular processes and can be imaged at nanoscale resolution using single-molecule localization microscopy (SMLM). Existing cluster analysis methods for regions of interest (ROIs) from SMLM data suffer from major limitations, such as unsuitability for heterogeneous datasets, failure to account for uncertainties in localization data, inability to analyze 3D

The dynamics and thermodynamics of the homodimer main protease protein SARS-CoV-2 are analysed at the level of Elastic Network Models (ENM). Correlated motions display a rich and heterogeneous dynamical structure, including allosterically correlated motions between the active sites in the two domains. Exhaustive 1-point and 2-point mutation maps, and their effect on binding free energies and allosteric

This technical study describes all-atom modeling and simulation of a fully-glycosylated full-length SARS-CoV-2 spike (S) protein in a viral membrane. First, starting from PDB:6VSB and 6VXX, full-length S protein structures were modeled using template-based modeling, de-novo protein structure prediction, and loop modeling techniques in GALAXY modeling suite. Then, using the recently-determined most

Many cellular processes require cell polarization to be maintained as the cell changes shape, grows or moves. Without feedback mechanisms relaying information about cell shape to the polarity molecular machinery, the coordination between cell polarization and morphogenesis, movement or growth would not be possible. Here we theoretically and computationally study the role of a genetically-encoded mechanical