- by Iqbal, K., Thiele, J. C., Saman, D., Peters, J. S., Thorpe, S., Tusk, S., Bardzil, J., Benesch, J. L. P., Kukura, P.Mass photometry (MP) is a powerful technique for studying biomolecular structure, interactions, and dynamics in solution. It detects and quantifies small reflectivity changes at a glass-water interface during protein (un)binding, with signals typically averaged over 100 milliseconds. However, particle motion at the point of single-molecule measurement can compromise key metrics such as mass resolution, sensitivity, and concentration. We present a three-dimensional convolutional residual network trained via supervised learning to classify landing events based on their spatiotemporal dynamics. By analysing 3D […]
- by Berg, B., Allard, J.A repeated pattern in cytoskeletal architecture is the aster, in which a number of F-actin filaments emerge star-shaped from a central node. Aster-based structures occur in cytoplasmic actin, the early stages of the cytokinetic ring in yeast, and in the context of biomimetic materials engineering. In this work, we use computational simulation to show that there is an optimal number of filaments per aster that maximizes rigidity, even at a fixed density of F-actin. This nonlinear dependence holds for both […]
- by Sugita, M., Noso, Y., Li, J., Fujie, T., Yanagisawa, K., Akiyama, Y.Improving membrane permeability is crucial in cyclic peptide drug discovery. Although the approach based on molecular dynamics (MD) simulation is widely used, it is computationally expensive. Alternatively, machine learning can predict membrane permeability at negligible cost, but it requires a larger dataset. There are only 7991 experimental values of membrane permeability available at the newly developed database. Another challenge in predicting membrane permeability using machine learning arises from the unique stable conformation of each cyclic peptide, which is strongly related […]
- by Walker, C., Mansour, D., Effiong, U., Jordan, D., Wang, L., Lafer, E. M., Alvarado, J., Belardi, B., Rangamani, P., Stachowiak, J. C., Rangamani, P.Protein condensates use multivalent binding and surface tension to assemble actin filaments into diverse architectures, reminiscent of filopodia and stress fibers. During this process, nucleation of new filaments and elongation of existing filaments inherently compete for a shared pool of actin monomers. Here we show that a balance between these competing processes is required to deform condensates of VASP, an actin binding protein, into structures with high aspect ratios. Addition of magnesium, which promotes filament nucleation, helped actin to deform […]
- by Yu, Z., Geng, J., Qiu, S., Zhang, W., He, M., Liu, X.Neurogranin (Ng) is enriched in the postsynaptic density, with mounting evidence of its importance to neural pathophysiology. Ng has long been recognized as a dedicated calmodulin (CaM)-binding/buffering protein. This study demonstrates that Ng, through its CaM-binding IQ domain, directly binds the distal C-terminus (DCT) of the L-type Ca2+ (CaV1) channel, as a de novo mechanism regulating excitation-transcription coupling. The Ng/DCT interaction attenuates autonomous C-terminus-mediated inhibition and promotes specific CaM delivery to CaV1, cooperatively enhancing channel gating (activation and inactivation), Ca2+ […]
- by Goldstein, R. E., Birwa, S. K., Yang, M.Nearly a decade ago it was discovered that the spherical cell body of the alga Chlamydomonas reinhardtii can act as a lens to concentrate incoming light onto the cell's membrane-bound photoreceptor and thereby affect phototaxis. Since many nearly transparent cells in marine environments have complex, often non-axisymmetric shapes, this observation raises fundamental, yet little-explored questions in biological optics about light refraction by the bodies of microorganisms. There are two distinct contexts for such questions: the absorption problem for incoming light, […]
- by Rienstra, N. M., Garvis, S., Sanchez, J. C., Sibert, B. S., Wright, E. R.We present a novel microfluidic flow cell, or Flow-enabled Light and Ultrastructural Imaging Device for Correlative Electron and Light Localization (FLUID-CELL), that connects fluorescence light microscopy (FLM) and cryo-electron microscopy (cryo-EM) for advanced biological imaging and ultrastructural and structural analyses. The design of the FLUID-CELL features a precisely engineered microchannel that maintains native cell culturing conditions, supporting correlation and enabling real-time observation by FLM, as well as subsequent cryo-EM analysis. In this study, this device enabled practical FLM imaging over […]
- by Losa, J., Simon, F., Linnik, D., Kaya, S. G., Stuart, M. C. A., Stetsenko, A., de Boer, R., Ho, F. Y., Incarnato, D., Stevens, J., van Eck, J., Fraaije, M. W., Weiss, L. E., van Teeffelen, S., Abeln, S., Guskov, A., Marrink, S.-J., Poolman, B., Heinemann, M.Molecular crowding in the bacterial cytoplasm restricts the diffusion of large molecules, impacting cellular processes. However, how nutrient availability influences cytoplasmic rheology is not well understood. With single-particle tracking in Escherichia coli, we observed a threefold variation in the diffusion of a 40-nm particle across exponential growth conditions. Previously suggested determinants of rheology did not account for this variation; instead, we found a strong anticorrelation between the diffusion coefficient and the abundance of amino acid metabolism proteins, persisting upon genetic […]
- by Munoz-Reyes, D., Fieseler, K. K., Winokan, M., Golding, M., Capkin, E., Ferla, M., Perez-Suarez, S., Tomlinson, C. W. E., Marples, P. G., Miro-Rodriguez, C., Aguado, L., Mansilla, A., Fearon, D., Thompson, W., von Delft, F., Sanchez-Barrena, M. J.Efficient drug discovery relies on workflows that integrate structural insights with rapid and cost effective exploration of chemical space. Here, we present a data-driven fragment-based lead discovery approach to target Neuronal Calcium Sensor 1 (NCS-1) protein-protein interactions (PPIs). This study represents the first implementation of a complete design-make-test-analyze cycle leading to the identification of micromolar affinity compounds with the potential to modulate NCS-1 interactions with key targets, including the G-protein chaperone Ric-8A and the dopamine D2 and cannabinoid CB1 receptors. […]
- by Favetta, B., Wang, H., Shi, Z., Schuster, B. S.Biomolecular condensates are protein-dense regions in cells that often arise from liquid-liquid phase separation. Interfacial tension is a key determinant of biomolecular condensate behavior, influencing condensate size and interactions with intracellular structures. Certain proteins and RNAs are known to selectively localize to the interface of condensates, where they can regulate condensate function in cells. Previously, we designed amphiphilic proteins that preferentially adsorb to the surface of condensates. These proteins contain one phase-separating domain (RGG) and one non-phase-separating domain (MBP or […]
- by Moffatt, L., Pierdominici-Sottile, G.Ligand-gated ion channels enable rapid cellular signaling by coupling extracellular cues to conformational transitions and ionic fluxes. ATP-activated P2X receptors, with their minimalist trimeric architecture, serve as models for studying allosteric activation. Although high-resolution structures reveal closed and open states, the physical basis of the "flip state" and the emergence of negative cooperativity remain unresolved. Here we combine a recursive Bayesian framework (MacroIR) applied to outside-out patch-clamp recordings, with molecular dynamics simulations, to show that P2X2 activation proceeds via a […]
- by Ricker, B., Dayan, N., Pelled, G., Gilad, A.Purpose: Investigation of the electromagnetic perceptive gene (EPG) protein and garnering evidence to suggest its use as a magnetogenetic tool for neuromodulation. Activation of EPG by electromagnetic field increases intracellular calcium levels, thus, we aimed to determine whether EPG influences the analogous manganese ion. This work yields potential for manganese-enhanced MRI (MEMRI) to be used to monitor EPG acting as a neuromodulator. Methods: HEK293FT cells expressing EPG were exposed to a MnCl2 solution and stimulated with a static or electromagnet. […]
- by Mansson, L. K., Warsaw, A. E., Wilbanks, E. G., Pitenis, A. A.Many hydrated and soft materials, including biological tissues and assemblies, permit fluid flow while retaining form and function. However, fundamental investigations of these complex materials are challenged by multiscale structural heterogeneity. Here, we characterized the structural and mechanical landscape of "pink berry" granular biofilms to determine the extent to which microscale heterogeneity influences macroscale material properties. We performed mechanical indentation measurements on intact granules as well as nanoindentation measurements on thin sections. We employed advanced imaging to correlate the internal […]
- by Foust, D. J., Kolli, D., Jessel, K., Hu, Z., Chapman, M. R., Biteen, J. S.Curli, which are the major proteinaceous component of the Escherichia coli biofilm extracellular matrix, help protect cells against environmental stressors including dehydration and antibiotics. Composed of the amyloid proteins CsgA and CsgB, curli self-assemble as these protomers are secreted into the extracellular space. The mechanisms of curli assembly and their functional roles within the extracellular matrix are incompletely understood. High-resolution imaging tools compatible with live-cell conditions provide a critical means to investigate the assembly and function of curli in their […]
- by Sarpangala, N., Gopinathan, A.Intracellular transport by teams of molecular motors is an essential cell-biological process that ensures the proper distribution of organelles, and other materials within cells. These teams of motors cooperate and compete in complex ways to achieve desired transport velocity and runlength. In-vitro experiments have observed that coupling motors through a lipid membrane that mimics in vivo membrane-bound cargoes leads to a higher cargo velocity. However, the mechanisms behind this increase in lipid cargo velocity are unclear. Here we seek to […]
- by Cuneo, M. J., Güllülü, O., Ammar, M.-R., Gui, X., Churion, K., Turk, M., O'Flynn, B., Sabri, N., Mittag, T.Proteostasis is critical for preventing oncogenesis. Both activating and inactivating mutations in the ubiquitin ligase subunit SPOP result in oncogenesis in different tissues. SPOP assembles into filaments that are multivalent for substrates, and substrates have multiple weak motifs for SPOP that are not activated via post-translational modifications. It is thus unclear how regulation is achieved. Here, we show that SPOP filaments circularize into rings that dimerize into up to 2.5 MDa-large, auto-inhibited double donuts. The equilibrium between double donuts and […]
- by Elsner, C., Roderer, D., Bordignon, E.The Bcl-2 protein Bcl-xL is a well-known inhibitor of apoptosis which can either directly inhibit the pore-forming Bcl-2 proteins, as Bax or Bak, or indirectly inhibit pore formation by sequestering the proapoptotic BH3-only proteins, as cBid or Bim. The structural basis of the inhibition of pore formation in the outer mitochondrial membrane by Bcl-xL is still largely unknown. Up to now, challenges in obtaining full-length Bcl-xL have hampered the full understanding of the inhibitory interactome at the membrane. Here we […]
- by Lee, J., Manon, V., Chen, J.In the host-pathogen arms race, herpesviruses and poxviruses encode proteins that sabotage the transporter associated with antigen processing (TAP), thereby suppressing MHC-I antigen presentation and enabling lifelong infection. Of the five known viral TAP inhibitors, only the herpes simplex virus protein ICP47 has been structurally resolved. We now report cryo-electron microscopy structures of TAP in complex with the remaining four: BNLF2a (Epstein-Barr virus), hUS6 (human cytomegalovirus), bUL49.5 (bovine herpesvirus 1), and CPXV012 (cowpox virus), assembling a structural atlas of viral […]
- by Mistryukova, L., Manuilov, V., Avchaciov, K., Fedichev, P. O.Designing small molecules that selectively bind to protein targets remains a central challenge in drug discovery. While recent generative models leverage 3D structural data to guide ligand generation, their applicability is limited by the sparsity and bias of structural resources. Here, we introduce ProtoBind-Diff, a structure-free masked diffusion model that conditions molecular generation directly on protein sequences via pre-trained language model embeddings. Trained on over one million active protein-ligand pairs from BindingDB, ProtoBind-Diff generates chemically valid, novel, and target-specific ligands […]
- by D'Urso, M., van den Bersselaar, P., Pragnere, S., Maiuri, P., Bouten, C. C. V., Kurniawan, N. A.During wound healing, fibroblasts undergo radical processes that impact their phenotype and behavior. They are activated, recruited to the injury site, assume a contractile phenotype, and secrete extracellular matrix proteins to orchestrate tissue repair. Thus, fibroblasts response require dynamic changes in cytoskeleton assembly and organization, adhesion morphology, and force generation. At the same time, fibroblasts experience changes in environmental stiffness during tissue wounding and healing. Although cells are generally known to use their adhesion contraction machinery to sense microenvironmental stiffness, […]
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