Dragnea Group

In a 2006 article entitled “The biological frontier”, Rob Phillips and Stephen Quake pointed out that material systems with characteristic dimensions between 1 and 100 nm experience an interesting confluence of energy scales for interactions which are traditionally associated with separate disciplines in physical sciences. Many biological systems are taking advantage of this confluence of scales as a principle for self-organization. Fascinating examples of synergies between mechanical, chemical, and thermodynamic driving forces include the self-assembly, packaging and release of nucleic acid by viruses. Understanding and emulating interaction synergies encountered in self-organizing biological matter has been until now the central theme of our research program. In terms of scope, our activities fit into three main categories:

1. Developing new tools and techniques that allow us to look further into the dynamics and organization of self-assembled biomolecular systems.
2. Fundamental work aiming at experimentally testing models concerned with the balance of forces driving molecular self-assembly.
3. Work dedicated to applications, until now mainly in the biomedical field, but currently exploring possibilities in energy transduction

Ongoing Research:

The ability of certain viruses to self-assemble spontaneously in vitro from nucleic acid and protein molecules has inspired us to re-direct the assembly process towards virus-like particles that package abiotic materials in a symmetric virus protein shell. Such constructs preserve some of the virus functions, which are carried by the virus protein shell, and in addition have physical properties coming form the encapsulated material. For example, virus-like particles carrying iron oxide or FePt cores will exhibit superparamagnetism and, at the same time, they will translocate in a plant host in similar ways to the wild type virus. We believe that in terms of applications, virus-like particles loaded with abiotic cargo offer new opportunities for therapeutics and diagnostic in nanomedicine due to unique characteristics which set them apart from other nanoparticle-based platforms: monodisperse size and surface properties, responsiveness to environmental cues often through dramatic conformational or dynamic changes, and availability of atomically-precise structural engineering through established molecular biology protocols. In terms of fundamentals, template virus-like particle assembly has significant potential in elucidating the relative importance of interactions between the different components of a virus since, for example, a virus capsid can be thought as a 2D crystal wrapped around a sphere and the amount and types of defects in the crystalline lattice depend on the spherical template diameter. Our long-term goals in continuing this research are the fulfillment of promises carried by the templated self-assembly of virus-like particles for applications in nanomedicine and fundamental understanding of virus-self assembly.

A research direction that we are currently pursuing and has actually stemmed from studies of nanoparticle-templated virus-like particles is the development of the photothermal imaging and single-particle tracking methodology. The technique can be applied in vivo or in vitro for studies of rare stochastic processes as a complement to in singulo fluorescence microscopy, which lacks the required dynamic range for such cases. With funding from NIH (NIGMS), we have shown that single virus-like particles can be imaged by photothermal microscopy in a turbid medium. Moreover, the large signal-to-noise ratio allows for clear discrimination between virus-protein coated particle cores and bare particle cores, which means that in the near future we should be able to measure the kinetics of uncoating for a single virus-like particle in real time.

Future new directions:

We have mentioned in the beginning a confluence of energies at nanoscale. Optical forces are part of this confluence as it was clearly illustrated with the help of optical tweezers in some of the most elegant experiments of in singulo biophysics. It has been pointed out in several theoretical papers that optical forces between nanoparticles illuminated by a focused laser beam at reasonable intensities should be strong enough to overcome Brownian motion and result in reversible particle association, and possibly crystallization. We believe that such optical forces could be a means of tuning chemical equilibrium in small nanoparticle solutions close to suprasaturation thus controlling crystallization of metallodielectric materials, which are of great technological interest but are difficult to manufacture, especially in 3D. Moreover, despite the technological importance of functionalized nanoparticles, there is a lack of direct experimental data concerning the effective pair potentials of small particles interacting in a solvent medium. Having an adjustable optical force between nanoparticles could in principle help to determine interparticle chemical forces. The short-term questions that we are currently attempting to answer are whether: 1) Optical forces between nanoparticles can be reliably measured free of thermal artifacts, preferably with a simple setup that probes an ensemble of particles rather than individual ones, and 2) Whether optical interactions can be used to reveal the differences in the surface treatments (leading to different pair potentials) that usually stabilize them in solution. These two questions are currently addressed in our lab using optical and electronic microscopy and EM simulations. We have preliminary data collected this summer which look promising. The setup is based upon an inverted microscope with phase contrast and a microfluidic microscopy chamber. The chamber channel is filled with a concentrated aqueous suspension of 10 nm diameter Au particles. The particles were functionalized by short polyethylene glycol chains. The PEG coating allows us to reach large particle concentrations, i.e. small interparticle distances, that are well beyond what citrate-stabilized colloids can provide. A green laser (5 mW) tuned to the surface plasmon resonance is focused on the sample and a white light phase contrast image recorded on a CCD after removing the laser through a holographic notch filter. Microscopy results suggest that the presence of the laser induces reversible concentration of the Au nanoparticles around the laser focus. We found that the profile of the nanoparticle distribution depends on the surface chemistry as well as on the laser intensity - which is what we are the most excited about. For example, carboxy-terminated PEG gives a noticeable different radial particle distribution than methoxy-terminated PEG (same nanoparticle size, concentration, and laser intensity). Data interpretation will require some modeling - Monte Carlo and MD simulations to study the interplay between the surface chemistry and the photoinduced-dipole interactions, a Maxwell-Garnett theory treatment to derive the dielectric constant change as a function of the particle distribution, and FDTD simulations to estimate the optical forces at work.

Here we seek to develop novel instrumentation for imaging biomolecular dynamics in real-time and at the scale of single biomolecular complexes that are much too small to be visualized by conventional optical microscopy. The proposed instrument is based on a simple wide-field optical microscope without any significant hardware modifications except for the sample support preparation, which sits at the heart of the optical super-resolution and requires standard microfabrication. The sample support is patterned similarly to a fly eye and its patterned domains serve as miniature far-field super-lenses. As a proof of principle, the release of the nucleic acid (RNA) payload from a single plant virus particle in solution will be observed for the first time with a spatial resolution of 20 nm and a time resolution of ~30 ms. Providing a general strategy that enables the observation of virus transformations leading to RNA presentation in physiological conditions will allow a better understanding of the molecular mechanisms leading to infection for a multitude of viruses, and could open new venues to antiviral therapeutics as well as novel approaches to bio-inspired targeted delivery of therapeutic and diagnostic agents.