PaperPlayer biorxiv biophysics

Link to bioRxiv paper:

http://biorxiv.org/cgi/content/short/2020.08.14.251223v1?rss=1

Authors: Rijal, K., Prasad, A., Das, D.

Abstract:

Protein thresholds have been shown to act as an ancient timekeeping device, such as in the time to lysis of E. coli infected with bacteriophage lambda. The time taken for protein levels to reach a particular threshold for the first time is defined as the first passage time of the protein synthesis system, which is a stochastic quantity. It had been shown previously that it was possible to obtain the mean and higher moments of the distribution of first passage times, but an analytical expression for the full distribution was not available. In this work, we derive an analytical expression for the first passage times for a long-lived protein. This expression allows us to calculate the full distribution not only for cases of no self-regulation, but also for both positive and negative self-regulation of the threshold protein. We show that the shape of the distribution matches previous experimental data on lambda-phage lysis time distributions. We study the noise in the precision of the first passage times by calculating the coefficient of variation (CV) of the distribution under various conditions. In agreement with previous results, we show that the CV of a protein that is not self-regulated is less than that of a self-regulated protein for both positive and negative regulation. We show that under conditions of positive self-regulation, the CV declines sharply and then roughly plateaus with increasing protein threshold, while under conditions of negative self-regulation, the CV declines steeply and then increases with increasing protein threshold. In the latter case therefore there is an optimal protein threshold that minimizes the noise in the first passage times. We also provide analytical expressions for the FPT distribution with non-zero degradation in Laplace space. These analytical distributions will have applications in understanding the stochastic dynamics of threshold determined processes in molecular and cell biology.

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Link to bioRxiv paper:

http://biorxiv.org/cgi/content/short/2020.08.14.251132v1?rss=1

Authors: Kaler, L., Iverson, E., Bader, S., Song, D., Scull, M. A., Duncan, G.

Abstract:

In this study, influenza A virus (IAV) and nanoparticle diffusion in human airway mucus was quantified using fluorescent video microscopy and multiple particle tracking. In previous work, it was determined that mucin-associated sialic acid acts as a decoy receptor for IAV hemagglutinin binding, and that virus passage through the mucus gel layer is facilitated through the sialic-acid cleaving enzyme, neuraminidase (NA), also present on the IAV envelope. However, our data suggests the mobility of IAV in mucus is significantly influenced by the mesh structure of the gel, as measured by nanoparticle probes, and NA activity is not required to facilitate virus passage through mucus gels. Using newly developed analyses, the binding affinity of IAV to the 3D mucus meshwork was estimated for individual virions with dissociation constants in the mM range, indicative of weak and reversible IAV-mucus interactions. We also found IAV diffusion significantly increased in mucus when treated with a mucolytic agent to break mucin-mucin disulfide bonds. In addition, IAV diffusion was significantly limited in a synthetic mucus model as crosslink density was systematically increased and network pore size was reduced. The results of this work provide important insights on how the balance of adhesive and physical barrier properties of mucus influence the dissemination of IAV within the lung microenvironment.

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Link to bioRxiv paper:

http://biorxiv.org/cgi/content/short/2020.08.14.250738v1?rss=1

Authors: Lu, H., Marti, J.

Abstract:

Ras is a family of related proteins participating in all animal cell lineages and organs which work as GDP-GTP binary switches and regulate cytoplasmic signalling networks able to control several cellular processes, playing an essential role in signal transduction pathways involved in cell growth, differentiation and survival. In this work, a G12D mutated farnesylated GTP bound KRas-4B protein has been simulated at the interface of a DOPC/DOPS/cholesterol model anionic cell membrane at the all-atom level. A specific long-lasting salt bridge connection between farnesyl and the hypervariable region of the protein has been identified as the main mechanism responsible of the binding of oncogenic farnesylated KRas-4B to the cell membrane, since this particular bond is absent in both wild-type and oncogenic methylated species of KRas-4B. This finding may lead to a deeper understanding of the mechanisms of protein binding and eventual growing and spreading inside cell membranes. From free energy landscapes obtained by well-tempered metadynamics simulations, we have characterised local and global minima of KRas-4B binding and revealed the main pathways between anchored and released states.

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Link to bioRxiv paper:

http://biorxiv.org/cgi/content/short/2020.08.12.248179v1?rss=1

Authors: Szulczewski, J. M., Inman, D. R., Proestaki, M., Notbohm, J., Burkel, B. M., Ponik, S. M.

Abstract:

Mechanosensory cues from the extracellular matrix underpin numerous cellular behaviors including tumor cell migration yet are influenced by the local structure and organization of the matrix in unknown ways. To investigate mechanical cues with respect to local collagen organization, we used a combination of intravital imaging of the mammary tumor microenvironment and 3D collagen gel systems with both migratory MDA-MB-231 cells and acellular pNIPAAm beads. We identified that fiber organization directs a bias in cell response along the axis of alignment. Using innovative methodology, we determined that local collagen alignment resulted in a 30-fold difference in directional cell-scale stiffness and also dramatically altered the rate at which cell-induced fiber displacements decayed over distance. Our results reveal differential mechanical properties across orthogonal directions in aligned matrices that provide sizeable cues to the cell and have important implications for cellular mechanosensing and cell-cell communication within the tissue microenvironment.

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Link to bioRxiv paper:

http://biorxiv.org/cgi/content/short/2020.08.12.247643v1?rss=1

Authors: Shashkova, S., Nyström, T., Leake, M. C., Wollman, A. J.

Abstract:

Most cells adapt to their environment by switching combinations of genes on and off through a complex interplay of transcription factor proteins (TFs). The mechanisms by which TFs respond to signals, move into the nucleus and find specific binding sites in target genes is still largely unknown. Single-molecule fluorescence microscopes, which can image single TFs in live cells, have begun to elucidate the problem. Here, we show that different environmental signals, in this case carbon sources, yield a unique single-molecule fluorescence pattern of foci of a key metabolic regulating transcription factor, Mig1, in the nucleus of the budding yeast, Saccharomyces cerevisiae. This pattern serves as a 'barcode' of the gene regulatory state of the cells which can be correlated with cell growth characteristics and other biological function.

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Link to bioRxiv paper:

http://biorxiv.org/cgi/content/short/2020.08.13.249862v1?rss=1

Authors: Sinnige, T., Meisl, G., Michaels, T. C. T., Vendruscolo, M., Knowles, T. P. J., Morimoto, R. I.

Abstract:

The accumulation of insoluble protein aggregates containing amyloid fibrils has been observed in many different human protein misfolding diseases, and their pathological features have been recapitulated in diverse model systems. In vitro kinetic studies have provided a quantitative understanding of how the fundamental molecular level processes of nucleation and growth lead to amyloid formation. 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 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 vitro and in cell culture, 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.

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Link to bioRxiv paper:

http://biorxiv.org/cgi/content/short/2020.08.13.249755v1?rss=1

Authors: Patel, D., Barnes, J. E., Davies, W. I. L., Stenkamp, D. L., Patel, J. S.

Abstract:

For many species, vision is one of the most important sensory modalities for mediating essential tasks that include navigation, predation and foraging, predator avoidance, and numerous social behaviors. The vertebrate visual process begins when photons of the light interact with rod and cone photoreceptors that are present in the neural retina. Vertebrate visual photopigments are housed within these photoreceptor cells and are sensitive to a wide range of wavelengths that peak within the light spectrum, the latter of which is a function of the type of chromophore used and how it interacts with specific amino acid residues found within the opsin protein sequence. Minor differences in the amino acid sequences of the opsins are known to lead to large differences in the spectral peak of absorbance (i.e. the {lambda} max value). In our prior studies, we developed a new approach that combined homology modeling and molecular dynamics simulations to gather structural information associated with chromophore conformation, then used it to generate statistical models for the accurate prediction of {lambda} max values for photopigments derived from Rh1 and Rh2 amino acid sequences. In the present study, we test our novel approach to predict the {lambda} max of phylogenetically distant Sws2 cone opsins. To build a model that can predict the {lambda} max using our approach presented in our prior studies, we selected a spectrally-diverse set of 11 teleost Sws2 photopigments for which both amino acid sequence information and experimentally measured {lambda} max values are known. The final first-order regression model, consisting of three terms associated with chromophore conformation, was sufficient to predict the {lambda} max of Sws2 photopigments with high accuracy. This study further highlights the breadth of our approach in reliably predicting {lambda} max values of Sws2 cone photopigments, evolutionary-more distant from template bovine RH1, and provided mechanistic insights into the role of known spectral tuning sites.

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Link to bioRxiv paper:

http://biorxiv.org/cgi/content/short/2020.08.11.245910v1?rss=1

Authors: Urbanska, M., Lüdecke, A., Walter, W. J., van Oijen, A. M., Duderstadt, K. E., Diez, S.

Abstract:

Optical trapping experiments have provided crucial insight into the operation of molecular motors. However, these experiments are mostly limited to one measurement at a time. Here, we describe an alternative, highly-parallel, microfluidics-based method that allows for rapid data collection. We applied tunable hydrodynamic forces to stepping kinesin-1 motors via DNA-tethered beads and utilized a large field-of-view to simultaneously track the velocities, run lengths and interaction times of hundreds of individual kinesin-1 molecules under varying resisting and assisting loads. Importantly, the 16-m long DNA tethers between the motors and the beads significantly reduced the vertical component of the applied force. Consequently, forces were predominantly exerted in the direction of motor movement, rather than away from the microtubule surface as unavoidable in conventional optical tweezers experiments. Our approach is readily applicable to other motors and constitutes a new methodology for parallelized single-molecule force studies.

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Link to bioRxiv paper:

http://biorxiv.org/cgi/content/short/2020.08.12.248500v1?rss=1

Authors: Kang, J. Y., Llewellyn, E., Chen, J., Olinares, P. D. B., Brewer, J., Chait, B. T., Campbell, E. A., Darst, S. A.

Abstract:

Transcription-coupled repair (TCR) is a sub-pathway of nucleotide excision repair (NER) that preferentially removes lesions from the template-strand (t-strand) that stall RNA polymerase (RNAP) elongation complexes (EC). Mfd mediates TCR in bacteria by removing the stalled RNAP concealing the lesion and recruiting Uvr(A)BC. We used cryo-electron microscopy to visualize Mfd engaging with a stalled EC and attempting to dislodge the RNAP. We visualized seven distinct Mfd-EC complexes in both ATP and ADP-bound states. The structures explain how Mfd is remodeled from its repressed conformation, how the UvrA-interacting surface of Mfd is hidden during most of the remodeling process to prevent premature engagement with the NER pathway, how Mfd alters the RNAP conformation to facilitate disassembly, and how Mfd forms a processive translocation complex after dislodging the RNAP. Our results reveal an elaborate mechanism for how Mfd kinetically discriminates paused from stalled ECs and disassembles stalled ECs to initiate TCR.

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Link to bioRxiv paper:

http://biorxiv.org/cgi/content/short/2020.08.12.248690v1?rss=1

Authors: Seitz, C., Casalino, L., Konecny, R., Huber, G., Amaro, R. E., McCammon, J. A.

Abstract:

Influenza neuraminidase is an important drug target. Glycans are present on neuraminidase, and are generally considered to inhibit antibody binding via their glycan shield. In this work we studied the effect of glycans on the binding kinetics of antiviral drugs to the influenza neuraminidase. We created all-atom in silico systems of influenza neuraminidase with experimentally-derived glycoprofiles consisting of four systems with different glycan conformations and one system without glycans. Using Brownian dynamics simulations, we observe a two- to eight-fold decrease in the rate of ligand binding to the primary binding site of neuraminidase due to the presence of glycans. These glycans are capable of covering much of the surface area of neuraminidase, and the ligand binding inhibition is derived from glycans sterically occluding the primary binding site on a neighboring monomer. Our work also indicates that drugs preferentially bind to the primary binding site (i.e. the active site) over the secondary binding site, and we propose a binding mechanism illustrating this. These results help illuminate the complex interplay between glycans and ligand binding on the influenza membrane protein neuraminidase.

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