Interaction of nano-TiO2 with lysozyme: insights into the enzyme toxicity of nanosized particles.
OBJECTIVE
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-TiO(2) 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.


METHODS
Laboratory-scale experiments were carried out to investigate the interactions of nano-TiO(2) with lysozyme. The interaction of nano-TiO(2) 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.


RESULTS
The interaction accorded with the Langmuir isothermal adsorption and the saturation number of lysozyme is determined to be 580 per nano-TiO(2) 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-TiO(2), the bacteriolysis activity of lysozyme was subjected to an obvious inhibition.


CONCLUSIONS
The two-step binding model of lysozyme was proposed, in which lysozyme was adsorbed on nano-TiO(2) particle surface by electrostatic interaction and then the hydrogen bond (N-H...O and O-H...O) formed between nano-TiO(2) 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 TiO(2) aggregate was formed in the presence of lysozyme where lysozyme may bridge between nano-TiO(2) particles. The coexistence of nano-TiO(2) 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.


CONCLUSIONS
Lysozyme was adsorbed on the nano-TiO(2) particle surface via electrostatic attraction and hydrogen bonds, and they also bridged among global nano-TiO(2) particles to form the colloidal particles. As a reasonable deduction of this study, nano-TiO(2) 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.


CONCLUSIONS
Our results strongly suggest that nano-TiO(2) has an obvious impact on biomolecules. Our data suggest that more attention should be paid to the potential toxicity of nano-TiO(2) 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.
Xu
Zhen
Z
State Key Laboratory of Pollution Control and Resource Reuse, College of Environmental Science and Engineering, Tongji University, Shanghai, 200092, China.
Liu
Xi-Wei
XW
Ma
Yin-Sheng
YS
Gao
Hong-Wen
HW
D006863:Hydrogen-Ion Concentration; D053768:Metal Nanoparticles / Q000737:chemistry / Q000633:toxicity* / Q000648:ultrastructure; D009113:Muramidase / Q000737:chemistry / Q000187:drug effects* / Q000648:ultrastructure; D009994:Osmolar Concentration; D011487:Protein Conformation / Q000187:drug effects; D017433:Protein Structure, Secondary / Q000187:drug effects; D013696:Temperature; D014025:Titanium / Q000737:chemistry / Q000633:toxicity*; D018675:Toxicity Tests
C009495:titanium dioxide; D014025:Titanium; D009113:Muramidase