Jasna Brujic

Direct observation of an ensemble of stable collapsed states in the mechanical folding of ubiquitin

Statistical theories of protein folding have long predicted plausible mechanisms for reducing the vast conformational space through distinct ensembles of structures. However, these predictions have remained untested by bulk techniques, because the conformational diversity of folding molecules has been experimentally unapproachable. Owing to recent advances in single molecule force-clamp spectroscopy, we are now able to probe the structure and dynamics of the small protein ubiquitin by measuring its length and mechanical stability during each stage of folding. Here, we discover that upon hydrophobic collapse, the protein rapidly selects a subset of minimum energy structures that are mechanically weak and essential precursors of the native fold. From this much reduced ensemble, the native state is acquired through a barrier-limited transition. Our results support the validity of statistical mechanics models in describing the folding of a small protein on biological timescales.

A ‘granocentric’ model for random packing of jammed emulsions

Packing problems are ubiquitous, ranging from oil extraction through porous rocks to grain storage in silos and the compaction of pharmaceutical powders into tablets. At a given density, particulate systems pack into a mechanically stable and amorphous jammed state. Previous theoretical studies have explored a connection between this jammed state and the glass transition, the thermodynamics of jamming and geometric modelling of random packings. Nevertheless, a simple underlying mechanism for the random assembly of athermal particles, analogous to crystalline ordering, remains unknown. Here we use three-dimensional measurements of packings of polydisperse emulsion droplets to build a simple statistical model in which the complexity of the global packing is distilled into a local stochastic process. From the perspective of a single particle, the packing problem is reduced to the random formation of nearest neighbours, followed by a choice of contacts among them. The two key parameters in the model—the available space around a particle and the ratio of contacts to neighbours—are directly obtained from experiments. We demonstrate that this ‘granocentric’ view captures the properties of the polydisperse emulsion packing—ranging from the microscopic distributions of nearest neighbours and contacts, to local density fluctuations, to the global packing density. Application of our results to monodisperse and bidisperse systems produces quantitative agreement with previously measured trends in global density. Our model therefore reveals a general principle of organization for random packing and may provide the foundations for a theory of jammed matter.

Force-dependent polymorphism in type IV pili reveals hidden epitopes

Through evolution, nature has produced exquisite nanometric structures, with features unrealized in the most advanced man-made devices. Type IV pili (Tfp) represent such a structure: 6-nm-wide retractable filamentous appendages found in many bacteria, including human pathogens. Whereas the structure of Neisseria gonorrhoeae Tfp has been defined by conventional structural techniques, it remains difficult to explain the wide spectrum of functions associated with Tfp. Here we uncover a previously undescribed force-induced quaternary structure of the N. gonorrhoeae Tfp. By using a combination of optical and magnetic tweezers, atomic force microscopy, and molecular combing to apply forces on purified Tfp, we demonstrate that Tfp subjected to approximately 100 pN of force will transition into a new conformation. The new structure is roughly 3 times longer and 40% narrower than the original structure. Upon release of the force, the Tfp fiber regains its original form, indicating a reversible transition. Equally important, we show that the force-induced conformation exposes hidden epitopes previously buried in the Tfp fiber. We postulate that this transition provides a means for N. gonorrhoeae to maintain attachment to its host while withstanding intermittent forces encountered in the environment. Our findings demonstrate the need to reassess our understanding of Tfp dynamics and functions. They could also explain the structural diversity of other helical polymers while presenting a unique mechanism for polymer elongation and exemplifying the extreme structural plasticity of biological polymers.

Measuring the distribution of interdroplet forces in a compressed emulsion system

The micromechanics of a variety of systems experiencing a structural arrest due to their high density could be unified by a thermodynamic framework governing their approach to ‘jammed’ configurations. The mechanism of supporting an applied stress through the microstructure of these highly packed materials is important in inferring the features responsible for the inhomo- geneous stress transmission and testing the universality for all jammed matter. In this paper, we present a novel method for measuring the force distribution within the bulk of a compressed emulsion system using confocal microscopy and explain our results with a simple theoretical model and computer simulations. We obtain an exponential distribution at large forces and a small peak at small forces, in agreement with previous experimental and simulation data for other particulate systems.

Biomimetic emulsions reveal the effect of mechanical forces on cell–cell adhesion

Cell–cell contacts in tissues are continuously subject to mechanical forces due to homeostatic pressure and active cytoskeleton dynamics. In the process of cellular adhesion, the molecular pathways are well characterized but the role of mechanics is less well understood. To isolate the role of pressure we present a dense packing of functionalized emulsion droplets in which surface interactions are tuned to mimic those of real cells. By visualizing the microstructure in 3D we find that a threshold compression force is necessary to overcome electrostatic repulsion and surface elasticity and establish protein-mediated adhesion. Varying the droplet interaction potential maps out a phase diagram for adhesion as a function of force and salt concentration. Remarkably, fitting the data with our theoretical model predicts binder concentrations in the adhesion areas that are similar to those found in real cells. Moreover, we quantify the dependence of the area of adhesion on the applied force and thus reveal adhesion strengthening with increasing external pressure even in the absence of active cellular processes. This biomimetic approach reveals a physical origin of pressure-sensitive adhesion and its strength across cell–cell junctions.

Specificity, Flexibility and Valence of DNA Bonds for Guided Emulsion Architecture

The specificity and thermal reversibility of DNA interactions have enabled the self-assembly of crystal structures, self-replicating materials and colloidal molecules. Grafting DNA onto liquid interfaces of emulsions leads to exciting new architectural possibilities due to the mobility of the DNA ligands and the patches they form between bound droplets. Here we show that the size and number of these adhesion patches (valency) can be controlled. Valence 2 leads to flexible polymers of emulsion droplets, while valence above 4 leads to rigid droplet networks. A simple thermodynamic model quantitatively describes the increase in the patch size with droplet radii, DNA concentration and the stiffness of the tether to the sticky-end. The patches are formed between droplets with complementary DNA strands or alternatively with complementary colloidal nanoparticles to mediate DNA binding between droplets. This emulsion system opens the route to directed self-assembly of more complex structures through distinct DNA bonds with varying strengths and controlled valence and flexibility.

Microscopic approach to the nonlinear elasticity of compressed emulsions.

Using confocal microscopy, we measure the packing geometry and interdroplet forces as a function of the osmotic pressure in a 3D emulsion system. We assume a harmonic interaction potential over a wide range of volume fractions and attribute the observed nonlinear elastic response of the pressure with density to the first corrections to the scaling laws of the microstructure away from the critical point. The bulk modulus depends on the excess contacts created under compression, which leads to the correction exponent α=1.5. Microscopically, the nonlinearities manifest themselves as a narrowing of the distribution of the pressure per particle as a function of the global pressure.

Attractive emulsion droplets probe the phase diagram of jammed granular matter.

It remains an open question whether statistical mechanics approaches apply to random packings of athermal particles. Although a jamming phase diagram has recently been proposed for hard spheres with varying friction, here we use a frictionless emulsion system in the presence of depletion forces to sample the available phase space of packing configurations. Using confocal microscopy, we access their packing microstructure and test the theoretical assumptions. As a function of attraction, our packing protocol under gravity leads to well-defined jammed structures in which global density initially increases above random close packing and subsequently decreases monotonically. Microscopically, the fluctuations in parameters describing each particle, such as the coordination number, number of neighbors, and local packing fraction, are for all attractions in excellent agreement with a local stochastic model, indicating that long-range correlations are not important. Furthermore, the distributions of local cell volumes can be collapsed onto a universal curve using the predicted k-gamma distribution, in which the shape parameter k is fixed by the polydispersity while the effect of attraction is captured by rescaling the average cell volume. Within the Edwards statistical mechanics framework, this result measures the decrease in compactivity with global density, which represents a direct experimental test of a jamming phase diagram in athermal systems. The success of these theoretical tools in describing yet another class of materials gives support to the much-debated statistical physics of jammed granular matter.

Kinetic control of the coverage of oil droplets by DNA-functionalized colloids

Controlled adsorption and phase behavior of colloids at liquid interface. We report a study of reversible adsorption of DNA-coated colloids on complementary functionalized oil droplets. We show that it is possible to control the surface coverage of oil droplets using colloidal particles by exploiting the fact that, during slow adsorption, compositional arrest takes place well before structural arrest occurs. As a consequence, we can prepare colloid-coated oil droplets with a “frozen” degree of loading but with fully ergodic colloidal dynamics on the droplets. We illustrate the equilibrium nature of the adsorbed colloidal phase by exploring the quasi–two-dimensional phase behavior of the adsorbed colloids under the influence of depletion interactions and present simulations of a simple model that illustrates the nature of the compositional arrest and the structural ergodicity.

Model for random packing of polydisperse frictionless spheres

We propose a statistical model for the random packing of frictionless polydisperse spheres in which the complexity of the global packing is distilled into a local stochastic process. We simplify the problem by considering the “granocentric” point of view of a single particle in the bulk, thereby reducing random packing to the assembly of nearest neighbours, followed by a random choice of contacts among them. The model is based on only two parameters, the available solid angle around each particle and the ratio of contacts to neighbors, which are both directly obtainable from experiments or simulations. As a result, the model analytically predicts the microscopic distributions of nearest neighbours and contacts, the local density fluctuations as well as the global density of the packing. We find that this granocentric view captures the essential properties of the polydisperse emulsion packing. This model suggests a general principle of organization for random packing and provides a statistical tool for quantifying the effect of the particle size distribution on the geometry of random packing in a variety of contexts of industrial relevance.