HANDY TOOL for nanoengineering: the Cowpea chlorotic mottle virus (CCMV) displays a soccer-ball pattern of the five-sided (turquoise) and six-sided (red and green) viral capsomeres in this computer image determined by x-ray crystallography. The capsomeres are composed of protein subunits with identical amino acid chains. In CCMV, there are three slightly different chain configurations or shapes of the proteins, represented by the three colors. Image: JACK JOHNSON, The Scripps Research Institute, and MARK YOUNG, Montana State University, Bozeman
A virus, essentially nucleic acid clothed in a protein coat, or capsid, is well designed for its lifestyle as a cellular parasite. Targeting, packaging and delivery have all been optimized over billions of years of evolution. To search out target cells, the viral coat incorporates recognition and docking sites for specific cell types. To stabilize its negatively charged genetic package, a virus may carry a remarkably high positive charge on the capsid interior. And once it arrives at its destination, a virus delivers its genes into the interior of the targeted cell, where it usurps cellular machinery for viral purposes. Now researchers are taking advantage of these viral systems to develop clever nanotechnology applications in medical imaging and drug delivery, as well as new approaches to building electronic devices (see sidebar: "Viral Nanoassemblers for Electronics").
Mark Young and Trevor Douglas, both at Montana State University, Bozeman, in conjunction with Jack Johnson¿s group at the Scripps Research Institute in La Jolla, Calif., spent a number of years sleuthing the structure and assembly of viruses. They focused on the well-studied Cowpea chlorotic mottle virus (CCMV). The viral coat of CCMV, like that of many viruses, is composed of identical protein subunits that self-assemble into a quasispherical shape known as an icosahedron. This geometry forms the largest volume of a given size that can be constituted from identical subunits, notes Young. The subunits are organized into five-sided and six-sided capsomeres, which are arranged to form a pattern similar to that on a soccer ball. CCMV has gated pores that open and close according to the chemistry of its surrounding environment.
Armed with an arsenal of CCMV knowledge, the researchers began to explore whether they could redesign the capsid to both incorporate an imaging agent and zoom in on new targets. In addition, they wondered, what could be packaged inside the viral capsid in place of nucleic acid? And how could the gates be triggered to make deliveries?
It turns out that the capsid, assembled without its nucleic acid and thus no longer infectious, can serve as a highly modifiable and versatile addition to the nanoengineer¿s toolbox. Conveniently, the empty capsid even self-assembles in the test tube or in yeast cells genetically engineered to produce subunits.
Visualizing the Threat
To conquer a metastasized cancer, physicians must identify the sites of new tumors and then selectively kill the wayward cells. CCMV capsids can potentially be engineered to achieve both goals.
For example, CCMV capsids could improve detection of tiny new tumors by magnetic resonance imaging (MRI). MRI identifies the differing responses of hydrogen atoms of water to the presence of a powerful magnetic field. Prior to being scanned, the patient may receive an injection of an imaging agent, most commonly gadolinium. The agent as currently given makes areas of interest more distinct but usually cannot resolve extremely small metastases.
Over the past two years, Young, Douglas and their colleagues significantly raised contrast levels in MRI images by incorporating the gadolinium atoms into CCMV protein shells. This promotes gadolinium¿s interaction with water molecules. That's because the gadolinium molecules--180 of which are woven into the 28-nanometer-diameter capsid--tend to be in higher concentration at any given location. Also, unlike today¿s gadolinium agent, which tends to clump, the gadolinium bound to the capsid surface keeps the atoms evenly distributed and available to interact with water.
To attach gadolinium to the capsids, the scientists exchanged the agent for the usual calcium--normally, during capsid assembly, calcium binds to the protein shell at sites between the subunits. To further knit gadolinium to the capsid, the researchers genetically engineered changes on the viral genome that optimized the binding sites for gadolinium.