Zhang Tong, Zhang Hao, Liu Guiliang, Gao Canzhu, Liu Rutao
School of Environmental Science and Engineering, China -America CRC for Environment & Health, Shandong Province, Shandong University, 27# Shanda South Road, Jinan, 250100, People's Republic of China.
J Fluoresc. 2014 Nov;24(6):1803-10. doi: 10.1007/s10895-014-1469-x. Epub 2014 Oct 17.
Heavy metals possess great endangerment to environment even human health because of their indissolubility and bioaccumulation. The toxicity of heavy metal ions (Cu(2+), Pb(2+), Zn(2+)) to trypsin was investigated by fluorescence, synchronous fluorescence, UV-vis absorption, circular dichroism (CD) spectroscopy, isothermal titration calorimetry (ITC), and enzyme activity assay. The experimental results showed that toxic effect of heavy metal ions was due to their own characteristic, rather than the electric charges of the ion. Zn(2+) could not show the obvious toxicity to trypsin, while the structure and function of trypsin was damaged when the enzyme explored to Cu(2+) and Pb(2+). From the spectra results, we found that Cu(2+) would bind with trypsin, which lead to the fluorescence quenched and hydrophobicity increased. Pb(2+) could also change the structure and reduce the activity of trypsin in high concentration. In vitro measurement, the toxicity order of heavy metal ions to trypsin is: Cu(2+) > Pb(2+) > Zn(2+). In addition, isothermal titration calorimetry analysis proved that the interactions between Cu(2+), Pb(2+), Zn(2+) and trypsin were all spontaneous and exothermic, which indicated the adverse effect of these heavy metal ions to trypsin.
重金属因其难溶性和生物累积性而对环境甚至人类健康具有极大危害。通过荧光光谱、同步荧光光谱、紫外可见吸收光谱、圆二色光谱（CD）、等温滴定量热法（ITC）和酶活性测定等方法研究了重金属离子（Cu(2+)、Pb(2+)、Zn(2+)）对胰蛋白酶的毒性。实验结果表明，重金属离子的毒性作用归因于其自身特性，而非离子的电荷。Zn(2+)对胰蛋白酶未表现出明显毒性，而当胰蛋白酶与Cu(2+)和Pb(2+)接触时，其结构和功能受到损害。从光谱结果来看，我们发现Cu(2+)会与胰蛋白酶结合，导致荧光猝灭且疏水性增加。Pb(2+)在高浓度时也会改变胰蛋白酶的结构并降低其活性。体外测定中，重金属离子对胰蛋白酶的毒性顺序为：Cu(2+)＞Pb(2+)＞Zn(2+)。此外，等温滴定量热法分析证明，Cu(2+)、Pb(2+)、Zn(2+)与胰蛋白酶之间的相互作用均为自发且放热的，这表明这些重金属离子对胰蛋白酶具有不利影响。
