Effects of nanomaterials on luciferase with significant protection and increased enzyme activity observed for zinc oxide nanomaterials.

This principle goal of this research was to examine the effects of various nanomaterials on the activity and behavior of the firefly enzyme luciferase. Nanomaterials have been found to stabilize, and in some instances, shown to increase the activity of enzymes. In this study gold, manganese oxide (MnO), and zinc oxide (ZnO) nanomaterials were utilized in order to test their effects on enzyme activity. Luciferase was used because its activity is easy to analyze, as it typically produces a large amount of bioluminescence easily detected by a Microtiter plate reader. Following incubation with the various nanomaterials, luciferase was subjected to degradation by several protein denaturing agents, such as heat, SDS, urea, ethanol, protease, hydrogen peroxide, and pH changes. Results indicated that luciferase activity is indeed affected when combined with nanomaterials, accompanied by both increases and decreases in enzyme activity depending on the type of nanomaterial and denaturing agent used. In most of the experiments, when incubated with ZnO nanomaterials, luciferase depicted significant increases in activity and bioluminescence. Additional experiments, in which human A375 cells were treated with luciferase-nanomaterial mixtures, also depicted increased enzyme activity and bioluminescence for luciferase incubated with ZnO nanomaterials. Ultimately, our findings indicated that when luciferase was subjected to multiple types of denaturation, zinc oxide nanomaterials dramatically preserved and increased enzyme activity and bioluminescence.

Interaction of MnO and ZnO nanomaterials with biomedically important proteins and cells.

Zinc and manganese nanomaterials may have potential for biomedical nanotechnology. Here first generation Zn and Mn oxide nanomaterials were prepared as determined by XRD. Transmission electron microscopy confirmed their nanoscale in two dimensions and revealed a rod or belt-like morphology for MnO or ZnO respectively. Association of MnO and ZnO to three model biomedically important proteins (albumin, protamine and thrombin) has been characterized by ultra-violet and dynamic laser light spectroscopy, UVS and DLLS respectively. UVS demonstrated a concentration-dependent loss of protein from the supernatant upon sedimentation of MnO or ZnO. Shifts in the surface charge of the MnO or ZnO by DLLS confirmed the protein's adsorption to the surface. MnO and ZnO were incubated with live human cells in culture (HeLa, A375 or 1321N1). A marked difference was observed for the two nanomaterials behavior in cell culture where the MnO could be discerned associating at the cell surface whereas the ZnO caused the cells to exhibit a rounded up morphology. Trypan blue dye exclusion studies demonstrated cytotoxicity of the ZnO at high concentrations 62.5-31.5 microg/mL whereas surprisingly the MnO demonstrated no cytotoxicity at any of the concentrations tested.