P. J. de Pablo

DNA-mediated anisotropic mechanical reinforcement of a virus

In this work, we provide evidence of a mechanism to reinforce the strength of an icosahedral virus by using its genomic DNA as a structural element. The mechanical properties of individual empty capsids and DNA-containing virions of the minute virus of mice are investigated by using atomic force microscopy. The stiffness of the empty capsid is found to be isotropic. Remarkably, the presence of the DNA inside the virion leads to an anisotropic reinforcement of the virus stiffness by ≈3%, 40%, and 140% along the fivefold, threefold, and twofold symmetry axes, respectively. A finite element model of the virus indicates that this anisotropic mechanical reinforcement is due to DNA stretches bound to 60 concavities of the capsid. These results, together with evidence of biologically relevant conformational rearrangements of the capsid around pores located at the fivefold symmetry axes, suggest that the bound DNA may reinforce the overall stiffness of the viral particle without canceling the conformational changes needed for its infectivity.

Jumping mode scanning force microscopy

In this letter, we present a new scanning probe microscopy mode, jumping mode, which allows the simultaneous measurement of the topography and of some other physical property of the sample. Essentially, at each image point first the topography of the sample is measured during a feedback phase of a cycle, and then the tip–sample interaction is evaluated in real time as the tip is moved away and towards the sample. Since the lateral motion is done out of contact the method is free, or nearly free, of shear forces. The general advantages of jumping mode are discussed. Finally, two different applications of this mode are presented. In addition to the topography, the first application measures the adhesion between the tip and the sample, while the second determines the corresponding electrostatic interaction.

A simple, reliable technique for making electrical contact to multiwalled carbon nanotubes

A simple method of making reliable electrical contact to multiwalled carbon nanotubes is described. With these contacts, current in the mA range can be routinely passed through individual multiwalled nanotubes without adverse consequences, thus allowing their resistance to be measured using a common multimeter. The contacts are robust enough to withstand temperature excursions between room temperature and 77 K. I(V) data from different multiwalled nanotubes are presented and analyzed.

Contactless experiments on individual DNA molecules show no evidence for molecular wire behavior

A fundamental requirement for a molecule to be considered a molecular wire (MW) is the ability to transport electrical charge with a reasonably low resistance. We have carried out two experiments that measure first, the charge transfer from an electrode to the molecule, and second, the dielectric response of the MW. The latter experiment requires no contacts to either end of the molecule. From our experiments we conclude that adsorbed individual DNA molecules have a resistivity similar to mica, glass, and silicon oxide substrates. Therefore adsorbed DNA is not a conductor, and it should not be considered as a viable candidate for MW applications. Parallel studies on other nanowires, including single-walled carbon nanotubes, showed conductivity as expected.

Monitoring dynamics of human adenovirus disassembly induced by mechanical fatigue

The standard pathway for virus infection of eukaryotic cells requires disassembly of the viral shell to facilitate release of the viral genome into the host cell. Here we use mechanical fatigue, well below rupture strength, to induce stepwise disruption of individual human adenovirus particles under physiological conditions, and simultaneously monitor disassembly in real time. Our data show the sequence of dismantling events in individual mature (infectious) and immature (noninfectious) virions, starting with consecutive release of vertex structures followed by capsid cracking and core exposure. Further, our experiments demonstrate that vertex resilience depends inextricably on maturation, and establish the relevance of penton vacancies as seeding loci for virus shell disruption. The mechanical fatigue disruption route recapitulates the adenovirus disassembly pathway in vivo, as well as the stability differences between mature and immature virions.

Minimizing tip-sample forces in jumping mode atomic force microscopy in liquid.

Control and minimization of tip-sample interaction forces are imperative tasks to maximize the performance of atomic force microscopy. In particular, when imaging soft biological matter in liquids, the cantilever dragging force prevents identification of the tip-sample mechanical contact, resulting in deleterious interaction with the specimen. In this work we present an improved jumping mode procedure that allows detecting the tip-sample contact with high accuracy, thus minimizing the scanning forces (-100 pN) during the approach cycles. To illustrate this method we report images of human adenovirus and T7 bacteriophage particles which are prone to uncontrolled modifications when using conventional jumping mode.

Built-in mechanical stress in viral shells.

Mechanical properties of biological molecular aggregates are essential to their function. A remarkable example are double-stranded DNA viruses such as the φ29 bacteriophage, that not only has to withstand pressures of tens of atmospheres exerted by the confined DNA, but also uses this stored elastic energy during DNA translocation into the host. Here we show that empty prolated φ29 bacteriophage proheads exhibit an intriguing anisotropic stiffness which behaves counterintuitively different from standard continuum elasticity predictions. By using atomic force microscopy, we find that the φ29 shells are approximately two-times stiffer along the short than along the long axis. This result can be attributed to the existence of a residual stress, a hypothesis that we confirm by coarse-grained simulations. This built-in stress of the virus prohead could be a strategy to provide extra mechanical strength to withstand the DNA compaction during and after packing and a variety of extracellular conditions, such as osmotic shocks or dehydration.

Scanning force microscopy jumping and tapping modes in liquids

In this work theoretical considerations of the performance of scanning force microscopy jumping mode and tapping mode in liquids are discussed. A priori, jumping mode should improve in a liquid environment compared to in air while the situation for tapping mode should become worse. In order to confirm this we present jumping and tapping mode images of DNA molecules absorbed on a mica substrate immersed in water. The experiments demonstrate that jumping mode is a suitable scanning force microscopy method by which to image soft samples in liquid and that it has similar or even better performance than those exhibited by tapping, but without the complex experimental requirements of this mode.

Scanning force microscopy three-dimensional modes applied to the study of the dielectric response of adsorbed DNA molecules

We have developed a set of working modes for scanning probe microscopy?(SPM), which generalizes the usual method of acquiring data. We call these modes three-dimensional?(3D) modes. Using these modes it is possible to measure typical SPM magnitudes, such as, for example, the tunnel current, the normal force and the amplitude or frequency of the cantilever oscillation, as a function of any other two magnitudes of the system: f(x1,x2). In this paper we present different examples of 3D modes. In particular, we have applied 3D modes to the study of the electrostatic interaction of co-adsorbed single walled carbon nanotubes and individual DNA molecules with a metallic scanning force microscopy tip. The data indicate that adsorbed DNA has a dielectric constant similar to that of the glass substrate.

Correlating the location of structural defects with the electrical failure of multiwalled carbon nanotubes

Electrical failure of carbon nanotubes was investigated by obtaining I(V) data with a voltage ramp from a rope of multiwalled carbon nanotubes. Noncontact scanning force microscope images were obtained before and after each I(V) curve until electrical failure of the tube resulted. Following this procedure, it was possible to correlate a defect on the surface of a nanotube with the exact location of the tube failure.