C&EN: TODAY'S HEADLINES - AEROGELS HOST PROTEIN ASSEMBLIES

STEVE RITTER, C&EN HOUSTON

In a new processing scheme developed by researchers at the Naval Research Laboratory (NRL) in Washington, D.C., cytochrome c molecules self-assemble into superstructures that can be incorporated into silica aerogels while retaining their ability to reversibly bind nitric oxide [Nano Lett., 3, 1463 (2003)].



SUPERSTRUCTURE Adsorbed layer of cytochrome c on a 10-nm gold nanoparticle provides a foundation for additional layers of the protein to self-assemble (schematic top). The partially denatured outer protein layer provides a protective shell. Micrograph (bottom) shows the gold-cytochrome c superstructure encapsulated in a highly porous silica aerogel. NAVAL RESEARCH LAB

Many bioactive molecules, proteins, and even whole cells have been successfully encapsulated in silica sol-gels with retention of their bioactivity, notes lead NRL researcher Debra R. Rolison. Until now, however, only hearty lipases that can withstand extreme conditions have been reported to survive further processing to form aerogels, the ultraporous, nearly light-as-air solids that result when residual solvent is removed from the pores of sol-gels.

While gas sensing is an important result of the work, "our real finding is what keeps the cytochrome c active during the high temperature and pressure necessary to supercritically process the liquid-filled sol-gel into an aerogel," Rolison says.

The researchers prepare cytochrome c superstructures by mixing the protein in buffered solution with colloidal gold. They find that a protein monolayer adsorbs on the gold nanoparticles, providing a foundation for additional protein layers to self-assemble--in all, about 10,000 protein molecules to one gold nanoparticle. The gold-protein composites are then added to a standard base-catalyzed silica sol just as the sol starts to gel, a process called "nanogluing." Rolison and her coworkers developed this process several years ago to incorporate metal oxides, carbon black, and other materials into aerogels.

Based on circular dichroism spectra, the researchers deduced that the outer protein layer of the composites is partially denatured during the sol-gel and supercritical fluid processing, providing a protective shell that allows the inner layers of cytochrome c molecules to escape unscathed. Without colloidal gold, supercritical processing results in disorganized, denatured proteins.

The NRL researchers monitor the integrity of the cytochrome c molecules throughout the processing steps by observing the 408-nm visible absorption (Soret band) associated with iron-heme binding in the folded protein. The intact protein superstructures in the aerogels also are visible in transmission electron micrographs.

The cytochrome c bioactivity was tested by passing NO over the aerogels. The researchers controlled reversible NO binding to the heme iron by toggling gas flow between argon and NO and observing the Soret band shift from 408 nm to 414 nm and back again.

Rolison and colleagues are unaware of other reports of colloid-directed assembly of protein superstructures. "We think such structures have been forming undetected in the presence of colloidal sols before now, even with nonheme proteins, but prior work with metal-colloid-adsorbed proteins included a centrifugation separation step in which shear stress may have destroyed the soft superstructures," she says.