Reactions Of C-terminal Metal Binding Domain Of Copper Transport Protein With Ag⁺and Hg²⁺

Through the research in recent years, people have found that metal ions are exist in the body of the organism broadly, not only consist in cytolemma and cytochylema, but also reside in protein and nucleic acid. These metal ions take part in energy conversion, material transfer, metabolic regulation, information transfer and other complex biochemical processes in vivo, and they play an important role in the growth and development of organisms. One of these important microelements is copper, it participates in all kinds of physiological processes as cofactor of various proteins and enzymes. After iron and zinc, copper occupies the third place in content during the transition elements in vivo. There have been many studies and explorations about the transport of copper ions in living organisms at home and abroad, especially, the transport mechanism of copper ions in the cell membrane.Studies find that the uptake of copper in human cells depends on Human Copper Transporter1(hCtrl) which belongs to the transmembrane transporter located in the cell membrane.It has been reported that hCtrl can transport some other metal ions besides copper ions, for instance, the combination and transport of zinc ions, iron ions and the extracellular N-terminal of hCtrl, and cisplatin transported into cells described through the researches reporting anticancer drugs. Selecting mercury ions (Hg2+) and silver ions (Ag+) which are severely harmful to human and other organisms and taking the octopeptide which lies in the intracellular C-terminal structure domain of hCtrl (C8:VDITEHCH) as the object of the study, this topic research goes into more details on the combination and the transport mechanism between these two metal ions and hCtrl by Nuclear magnetic resonance (NMR), Electrospray ionization mass spectrometry (ESI-MS), Ultraviolet and visible spectrum (UV-vis) and other experiment measures.Chapter1mainly discusses the research background, development and work meaning of hCtrl and gives a brief introduction to the effect that Hg2+and Ag+have on human and other organisms. Section1mainly introduces the background and meaning of the research on hCtrl through literature. Section2mainly introduces the structure of hCtrl and the function of hCtrl in the human body. Section3mainly introduces how copper is transported from the outside to the inside of the cell in detail and illustrates the mechanism of Cu (Ⅰ) transported by hCtrl. Section4mainly introduces that the effect of Hg2+and Ag+having on human and other organisms, the advantages and disadvantages of Hg2+and Ag+, and the elimination of toxicity of the two metal ions.Chapter2studies the reaction between Hg2+and the C-terminal octapeptide of hCtrl. Selecting the C-terminal octapeptide of hCtrl for the research, the experiment detects and analyzes the reaction of Hg2+and C8by NMR, ESI-MS, UV-vis and other experiment measures. The NMR experiment data shows that the Histidine (His) can coordinate with Hg2+. In the electrospray ionization mass spectrum (ESI-MS), the spectral peak of C8binding to Hg2+in a mole ratio of2:1explains that C8binds to Hg2+by the way of dimer form of bridging, while the data of the tandem mass spectrometry (MS/MS) shows that the amino acid residues of Hg2+binding to C8may be the HCH motif. Data of UV states shows that the sulfhydryl group (-SH) of cysteine (Cys) of C8can quickly bind to Hg2+in the mole ratio of2:1and the binding constant isn’t small, and this conclusion confirms the results of MS.Chapter3studies the reaction between Ag+and the C-terminal octapeptide of hCtrl. In the experimental process, the peptide of C8, the experimental considerations and the experiment measures are the same as the experiment of Hg2+, only pH is directly transferred to neutral replacing the phosphate buffer (it can precipitate Ag+) in the NMR experiment. Data of NMR reveals that His also binds to Ag+. The spectral peak of C8binding to Ag+in a mole ratio of1:1of MS confirms the results of UV, and the data of MS/MS illustrates that Ag+not only binds to HCH but also binds to other amino acids far apart. Data of UV reveals that the sulfhydryl group of Cys of C8can bind to Ag+in the mole ratio of1:1, and the combination of low constant explains that Ag+binds to C8at a moderate speed.