Interaction of nano-TiO2 with lysozyme: insights into the enzyme toxicity of nanosized particles

Nanomaterials have been used increasingly in industrial production and daily life, but their human exposure may cause health risks. The interactions of nanomaterial with functional biomolecules are often applied as a precondition for its cytotoxicity and organ toxicity where various proteins have been investigated in the past years. In the present study, nano-TiO2 was selected as the representative of nanomaterials and lysozyme as a representative for enzymes. By investigating their interaction by various instrumentations, the objective is to identify the action sites and types, estimate the effect on the enzyme structure and activity, and reveal the toxicity mechanism of nanomaterial. Laboratory-scale experiments were carried out to investigate the interactions of nano-TiO2 with lysozyme. The interaction of nano-TiO2 particles with lysozyme has been studied in the analogous physiological media in detail by UV spectrometry, fluorophotometry, circular dichroism (CD), scanning electron microscope, zeta-potential, and laser particle size. The interaction accorded with the Langmuir isothermal adsorption and the saturation number of lysozyme is determined to be 580 per nano-TiO2 particle (60 nm of size) with 4.7 x 10(6) M-1 of the stability constant in the physiological media. The acidity and ion strength of the media obviously affected the binding of lysozyme. The warping and deformation of the lysozyme bridging were demonstrated by the conversion of its spatial structure from alpha-helix into a beta-sheet, measured by CD. In the presence of nano-TiO2, the bacteriolysis activity of lysozyme was subjected to an obvious inhibition. The two-step binding model of lysozyme was proposed, in which lysozyme was adsorbed on nano-TiO2 particle surface by electrostatic interaction and then the hydrogen bond (N-H center dot center dot center dot O and O-H center dot center dot center dot O) formed between nano-TiO2 particle and polar side groups of lysozyme. The adsorption of lysozyme obeyed the Langmuir isothermal model. The binding of lysozyme is dependent on the acidity and ion strength of the media. The bigger TiO2 aggregate was formed in the presence of lysozyme where lysozyme may bridge between nano-TiO2 particles. The coexistence of nano-TiO2 particles resulted in the transition of lysozyme conformation from an alpha-helix into a beta-sheet and a substantial inactivation of lysozyme. The beta-sheet can induce the formation of amyloid fibrils, a process which plays a major role in pathology. Lysozyme was adsorbed on the nano-TiO2 particle surface via electrostatic attraction and hydrogen bonds, and they also bridged among global nano-TiO2 particles to form the colloidal particles. As a reasonable deduction of this study, nano-TiO2 might have some toxic impacts on biomolecules. Our data suggest that careful attention be paid to the interaction of protein and nanomaterials. This could contribute to nanomaterial toxicity assessment. Our results strongly suggest that nano-TiO2 has an obvious impact on biomolecules. Our data suggest that more attention should be paid to the potential toxicity of nano-TiO2 on biomolecules. Further research into the toxicity of nanosized particles needs to be carried out prior to their cell toxicity and tissue toxicity. These investigations might serve as the basis for determining the toxicity and application of nanomaterials.