All tumor volumes in the five groups increased gradually during follow-up

sociated with the higher catalytic activity. Previously, based on a systematic mutagenesis study, we have proposed a nano-channel to be responsible for relaying the mutation effect from the extra domain to the catalytic machinery, which are mainly constituted by residues next to S284-T285-I286 on the extra domains as well as N-finger residues on the catalytic folds. Because the mutation of STI to Ala with a small side chain results in a cavity, in STI/A the two extra domains come slightly closer as described above, and consequently the volume of the nano-channel is 1293 A3 in STI/A, slightly smaller than that in WT. Molecular dynamics simulations Molecular dynamics simulation is a powerful tool to provide insights into the dynamic mechanism that underlies protein function. Therefore, we conducted 100-ns MD simulations for the wild-type and our present STI/ A structures, both of which have been determined at pH 6.0 and in crystals of the space group C2. 5 Dynamical Enhancement of SARS-CoV 3CLpro larger overall RMSD than WT, with average RMSD purchase HC030031 values over 100-ns simulations of 2.6460.32, 1.9060.26 and 2.6860.42 A for three separate simulations of STI/A; and 1.6060.26, 1.6660.31 and 1.6360.25 A for three simulations of WT. However, the averaged Ca RMSD values of the chymotrypsin folds of STI/A are only slightly larger than those of WT, with average RMSD values over 100-ns simula tions of 1.4060.21, 1.2060.16 and 1.4460.26 A for three simulations of STI/A; and of 1.2860.16, 1.1760.17 and 1.2560.15 A for three simulations of WT. On the other hand, the extra domains of STI/A have dramatically higher fluctuations than WT in simulations as indicated by its higher RMSD values, with average RMSD values over 100-ns simula tions of 3.5760.54, 2.1960.53 and 3.2660.86 A for three simulations of STI/A and of 1.7560.51, 1.6260.31 and 1.6160.31 A for three simulations of WT. These results clearly demonstrate that the larger RMSD values for the dimeric STI/A are mostly resulting from the motions of the extra domains. This premise is further evident from the root-mean-square fluctuation averaged over 100 ns for both STI/A and WT residues. Indeed, for STI/A, the distance d and angle H as defined in 6 Dynamical Enhancement of SARS-CoV 3CLpro We have also calculated the hydrogen bond occupancy of all simulations for both STI/A and WT, and Dynamic behavior of the catalytic dyad and oxyanion hole In the previously-determined crystal structures of SARS 3CLpro, the distance between NE2 of His41 and SG of Cys145 ranges from 3.6 to 3.9 A. Furthermore, all previous MD simulations revealed that the dynamic stability of this distance is extremely critical for the stable formation of a hydrogen bond, which appears to be pivotal for maintaining the catalytic competency of the SARS 3CLpro. Also for the active WT enzyme, this distance has been previously demonstrated to be dynamically stable in MD simulations. Indeed, as in our previous simulations, the distances of both PubMed ID:http://www.ncbi.nlm.nih.gov/pubmed/19655565 protomers of the WT were found to be dynamically stable for all 30 ns while those of the inactive N214A mutant became unstable and larger despite sharing a superimposable crystal structure to the WT. In our current MD simulations of 100 ns, the distances between NE2 of His41 and SG of Cys145 of STI/A are: 3.8060.60 A, 3.9160.51 and 3.7860.50 A respectively for three simulations of the protomer 1, and 3.9160.48, 3.8760.45 and 3.8660.48 A respectively for the protomer 2. In contrast, the distances in WT are