## 3 Discussion
### The Structure Analysis of Platinum Agents Combining with the Monomeric and Dimeric Atox1 Protein
To further investigate the protein unfolding induced by the platinum agents binding, the CASTp program was employed to calculate the size and shape of platinum agents binding site pockets. Such analysis was able to provide a comprehensive and detailed quantitative characterization of protein surface pockets and protein deformation [38, 39, 40]. It can be seen from Figure 4 that the binding site pocket sizes are 0 A\({}^{2}\)/0 A\({}^{3}\), 60.3 A\({}^{2}\)/45.5 A\({}^{3}\), 44.9 A\({}^{2}\)/32.2 A\({}^{3}\) and 82.6 A\({}^{2}\)/61.5 A\({}^{3}\) for the apo-Atox1, CisPt + Atox1, TransPt + Atox1 and OxaliPt + Atox1 models, respectively, which presented that the platinum agents binding to the Atox1 protein cause the formation of the platinum agents binding pocket around the residues Val8, Asp9, Cys12, Gly13, Gly14 and Cys15 _etc._ in the \(\beta\)1-\(\alpha\)1 loop of the protein, consequently the deformation of the original Atox1 structure. Specially, the large ligand size of oxaliplatin agent plays a key role in the formation of a largest binding pocket in the Atox1 protein. Summarily, based on the analysis for the unfolding residues and the pocket sizes around the binding sites of Atox1, the order of the extent of the Atox1 unfolding is the oxaliplatin, cisplatin and transplatin agents, which is consistent with the previous experimental result reported by [PERSON] and co-workers [29]. In particular, the oxaliplatin agent binding to the Atox1 protein is able to cause the larger deformation of the protein with the formation of large binding pockets and the more extensions of the interhelical angle of \(\alpha\)1 and \(\alpha\)2 helices, and the increase of the centroid distances of the \(\beta\)1 strand and \(\alpha\)1 helix, compared with other two cisplatin and transplatin agents. Furthermore, the large deformation of the Atox1 protein induced by the oxaliplatin agent binding favors the affinity of the platinum agent to the protein. Moreover, due to Atox1 functionally acting as a Cu transport protein, it is possible that Cu and Pt bind to two Cys residues in the Atox1 protein simultaneously, which was showed _in vitro_ as reported previously [25]. It is suggested that a Cu ion and a platinum agent simultaneously binding to two Cys residues may cause the more looseness of the binding loop region of Atox1 due to the appearance of an additional Cu ion in the Atox1 protein, resulting in the large deformation of the Atox1 protein. Based on the experimental studies of the structure of a ternary cisplatin + 2 Atox1 complex, the two Atox1 proteins are linked by the Pt(II) center in the asymmetric unit with two \(\alpha\)-helical layers at the inside of the complex and two \(\beta\)-sheet layers at the outside [23]. It can be seen from the average structures in Figure 8 that the ligand plane of the cisplatin agent in the CisPt + 2 Atox1 model almost parallels to the two \(\alpha\) helical layers of the Atox1 dimer, and is far away from the Atox1 protein-Atox1  
Figure 9: **(a)** The change of the interhelical angle (\(\Delta\theta\)) between the \(\alpha\)1 and \(\alpha\)2 helices for the apo-Atox1 model (**light blue**) and the binary models (**cyan**); and **(b)** The time-dependence of the interhelical angles along with the respective integrated distributions for the apo-Atox1 (**light blue**), CisPt + Atox1 (**green**), TransPt + Atox1 (**magenta**) and OxaliPt + Atox1 (**cyan**) models. protein interface towards the outside of the Atox1 dimer. For the OxaliPt + 2 Atox1 model, the diaminocyclohexane plane of the oxaliplatin agent is perpendicular to the two \(\alpha\) helical layers, and is also far away from the Atox1 protein-Atox1 protein interface towards the outside of the Atox1 dimer. Such conformation with the ligand plane towards the outside of the Atox1 dimer for the two models results in the insignificant interaction between the platinum ligand and the Atox1 dimer.