In this work, the interaction between nano-TiO2 and trypsin was investigated, and the mechanisms of the interaction were explored by the methods of UV–vis detection, circular dichroism (CD), and fluorescence. The results clearly demonstrated that nano-TiO2 had an inhibitory effect on the enzyme activity. The activity was decreased to 64% of the untreated trypsin in the presence of 300 μg/ml nano-TiO2. UV spectrometry proved that nano-TiO2 had a strong physical absorption effect on trypsin, and the CD spectra revealed that the secondary structure of trypsin was partly destroyed while bound together with nano-TiO2. The ratio of α-helix increased from 7.9% to 12.8% in the presence of 100 μg/ml TiO2 while the ratio of β-sheet decreased from 48.7% to 36.4%. Furthermore, the fluorescence spectrometry indicated that nano-TiO2 could quench the intrinsic fluorescence of trypsin through static quenching. Meanwhile, the binding constant was calculated to be 1, and the process of binding of trypsin on nano-TiO2 was a spontaneous molecular interaction procedure in which electrostatic interaction plays a major role. Our study was to provide a useful approach for evaluating the health risk of nanomaterials on level of proteins.
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The work was financially supported by the 973, 863 project of the Ministry of Science and Technology (No.2010CB912604, 2010CB933901, 2007AA022004), Research Fund for the Doctoral Program of Higher Education of China (Grant no.20090072120019), Shanghai Educational Development Foundation (Grant no. 2008CG26).
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