Enzyme Stabilization and Immobilization

Amanda S. Harper-Leatherman, Jean Marie Wallace, Debra R. Rolison, Harper-Leatherman, Amanda S., Wallace, Jean Marie, Rolison, Debra R.

Sol-gel-derived aerogels are three-dimensional, nanoscale materials that combine large surface area with high porosity. These traits make them useful for any rate-critical chemical process, particularly sensing or electrochemical applications, once physical or chemical moieties are incorporated into the gels to add their functionality to the ultraporous scaffold. Incorporating biomolecules into aerogels, other than such rugged species as lipases or cellulose, has been challenging due to the inability of most biomolecules to remain structurally intact within the gels during the necessary supercritical fluid (SCF) processing. However, the heme protein cytochrome c (cyt.c) forms self-organized superstructures around gold (or silver) nanoparticles in buffer that can be encapsulated into wet gels as the sol undergoes gelation. The guest-host wet gel can then be processed to form composite aerogels in which cyt.c retains its characteristic visible absorption. The gold (or silver) nanoparticle-nucleated superstructures protect the majority of the protein from the harsh physicochemical conditions necessary to form an aerogel. The Au~cyt.c superstructures exhibit rapid gas-phase recognition of nitric oxide (NO) within the bioaerogel matrix, as facilitated by the high-quality pore structure of the aerogel, while remaining viable for weeks at room temperature. More recently, careful control of synthetic parameters (e.g., buffer concentration, protein concentration, SCF extraction rate) have allowed for the preparation of cyt.c-silica aerogels, sans nucleating nanoparticles; these bioaerogels also exhibit rapid gas-phase sensing while retaining protein structural stability.