Abstract Uropathogenic Escherichia coli attach to tissues using pili type 1. Each pilus is composed by thousands of coiled FimA domains followed the tip fibrillum, FimF-FimG-FimH. The are linked non-covalent β-strands that must resist mechanical forces during attachment. Here, we use single-molecule force spectroscopy measure contribution each domain stability and monitor oxidative folding mechanism a single Fim assisted periplasmic FimC oxidoreductase DsbA. We demonstrate bear high...

Significance Talin is an adaptor protein that transduces mechanical signals into biochemical cues by recruiting a network of ligands in force-dependent way. These force have complex nature, oscillate time with different frequency components, and are often embedded noise. However, most assays to explore the mechanics force-sensing proteins rely on simple perturbations, such as constant or ramped forces. Here, we use our magnetic tweezers design subject single talin domains oscillatory forces...

Mechanical force regulates talin-vinculin association and defines an optimal binding range.

Surface sensing initiates bacterial colonization of substrates. The protein PilY1 plays key roles during this process—surface detection, host adhesion, and motility—while experiencing mechanical perturbations varying magnitudes. In Pseudomonas aeruginosa , the adhesion motility functions are associated with integrin calcium ligand-binding sites; however, how forces influence PilY1’s dynamics its interactions these ligands remain unknown. Here, using single-molecule magnetic tweezers, we...

FtsZ filaments participate in bacterial cell division, but it is still not clear how their dynamic polymerization and shape exert force on the underlying membrane. We present a theoretical description of individual that incorporates information from molecular simulations. The structure crystallized Methanococcus jannaschii dimer was used to model pentamer showed curvature twist. estimated bending torsion angles between monomers fluctuations were included description. MD data also permitted...

Bacteria anchor to their host cells through adhesive pili, which must resist the large mechanical stresses induced by as it attempts dislodge pathogens. The pili of gram-positive bacteria are constructed a single polypeptide made hundreds pilin repeats, contain intramolecular isopeptide bonds strategically located in structure prevent unfolding under force, protecting pilus from degradation extant proteases and oxygen radicals. Here, we demonstrate design short peptide that blocks formation...

Cell-surface proteins are central for the interaction of cells with their surroundings and also associated numerous diseases. These molecules exposed to mechanical forces, but exact relation between force functions pathologies cell-surface is unclear. An important protein CD4, primary receptor HIV-1. Here we show that activates conformational chemical changes on CD4 may be during viral attachment. We have used single-molecule spectroscopy analysis HIV-1 infectivity demonstrate extension...

For the first time, respiratory complex I has been reconstituted on an electrode preserving its structure and activity. Respiratory is a membrane-bound enzyme that essential function in cellular energy production. It couples NADH:quinone oxidoreduction to translocation of ions across (in prokaryotes) or mitochondrial membranes. Therefore, contributes establishment maintenance transmembrane difference electrochemical potential required for adenosine triphosphate synthesis, transport,...

The pathogenic bacterium Yersina pestis is protected from macrophage engulfment by a capsule like antigen, F1, formed of long polymers the monomer protein, Caf1. However, despite importance this pathogen, mechanism protection was not understood. Here we demonstrate how F1 protects bacteria phagocytosis. First, show that Escherichia coli expressing showed greatly reduced adherence to macrophages. Furthermore, few cells did adhere remained on surface and were engulfed. We then inserted,...

FtsZ is a bacterial cytoskeletal protein involved in cell division. It forms ringlike structure that attaches to the membrane complete binds and hydrolyzes GTP, assembling into polymers GTP-dependent manner. To test how orientation of monomers affects curvature filaments on surface, we performed site-directed mutagenesis E. coli insert cysteine residues at lateral locations orient planar lipid bilayers. The E93C S255C mutants were overproduced, purified, found be functionally active...

Proteins that operate under force—cell adhesion, mechanosensing—exhibit a wide range of mechanostabilities. Single-molecule magnetic tweezers has enabled the exploration dynamics force these proteins with subpiconewton resolution and unbeatable stability in 0.1-120 pN range. However, featuring high mechanostability (>120 pN) have remained elusive this technique been addressed Atomic Force Microscopy (AFM), which can reach higher forces but displays less resolution. Herein, we develop...

Abstract The classical “one sequence, one structure, function” paradigm has shaped much of our intuition how proteins work inside the cell. Partially due to insight provided by bulk biochemical assays, individual biomolecules are assumed behave as identical entities, and their characterization relies on ensemble averages that flatten any conformational diversity into a unique phenotype. While emergence single-molecule techniques opened gates interrogating molecules, technical shortcomings...

Gram positive bacteria colonize mucosal tissues against large mechanical perturbations, such as coughing, which generate shear forces that exceed the ability of non-covalent bonds to remain attached. To overcome these challenges, pathogen Streptococcus pyogenes utilizes protein Cpa, a pilus tip-end adhesin equipped with Cys-Gln thioester bond. The reactivity this bond towards host surface ligands enables covalent anchoring bacterium, allowing it resist shocks; however, colonization also...

Cells continually sample their mechanical environment using exquisite force sensors such as talin, whose folding status triggers mechanotransduction pathways by recruiting binding partners. Mechanical signals in biology change quickly over time and are often embedded noise; however, the mechanics of force-sensing proteins have only been tested simple protocols, constant or ramped forces. Here, our magnetic tape head tweezers design, we measure dynamics single talin response to external noise...

Vinculin binds unfolded talin domains in focal adhesions, which recruits actin filaments to rein-force the mechanical coupling of this organelle. However, mechanism by interaction is regulated, and its impact force transmission properties mechanotransduction pathway remain unknown. Here, we use magnetic tweezers spectroscopy measure binding vinculin head R3 domain under physiological loads. For first time, resolve individual events as a short contraction polypeptide due reformation helices...

ABSTRACT Bacteria anchor to their host cells through adhesive pili, which must resist the large mechanical stresses induced by as it attempts dislodge pathogens. The pili of Gram-positive bacteria are constructed a single polypeptide made hundreds pilin repeats, contain intramolecular isopeptide bonds strategically located in structure prevent unfolding under force, protecting pilus from degradation extant proteases and oxygen radicals. Here, we demonstrate design short peptide that blocks...