Structural Features and Dynamics Study of Viral Channel Forming Proteins in the Lipid Bilayer using Molecular Dynamics Simulation Method

• Main Authors: MONOJ MON KALITA, 柯孟明
• Other Authors: WOLFGANG B. FISCHER
• Format: Others
• Language: en_US
• Published: 2016
• Online Access: http://ndltd.ncl.edu.tw/handle/46562730166701107590

博士 === 國立陽明大學 === 生醫光電研究所 === 104 === Utilizing the combination of powerful MD techniques and a fine-grained search protocol which search the conformational space consisting of distance, rotational angle, absolute angle, and tilt, the first part of this study focuses on developing models for monomeric as well as a hexameric bundle of p7 protein from Hepatitis C Virus (HCV). It is a 63 amino acid polytopic membrane protein with two transmembrane domains (TMDs) and is involved in the modulation of electrochemical gradients across membranes within infected cells. Although there is a debate on the symmetry of the bundle architecture, till date only a single structure of p7 is available in the database, which is hexameric in symmetry and is based on solution NMR experiment. Using evidence and predictions as mentioned above, an optimized protocol is proposed to generate in silico model of p7. The in silico model is the result of combination of protein-protein docking method under MOE platform and MD simulation. This model is similar to one predicted by Patargias et al. (1) in the context of the symmetry and position of His-17, however the great difference between the two is the length of the helical motifs. Full length helical motifs are taken into consideration in this study while it is not the case in the latter. Genotype-specific structural features of p7 have been investigated for the in silico model (genotype 1a) as well as the recently reported experimental monomeric and hexameric structures (genotype 1b and 5a respectively). In addition to that, comparison of their channel gating behavior has been simulated. All the bundles tend to turn into a compact structure during MD simulations in hydrated lipid bilayers, as well as when simulated at ‘low pH’, which is proposed to trigger channel opening. Both the genotype 1a and 1b channel models are gated via movement of the parallel aligned helices, but the scenario for the genotype 5a protein is more complex, with a short N-terminal helix being involved. However, all bundles display pulsatile dynamics identified by monitoring water dynamics within the pore. In the second part, dynamics of the generated model of hexameric p7 protein of genotype 1a is studied using sophisticated trajectory analysis method such as full correlation analysis (FCA) along with the experimentally established structure of genotype 5a. FCA is useful in identifying the correlated principal dynamics in a protein. Results of FCA reveals that though structural differences are there in both bundles they screen local minima during the simulation with the asymmetric collective motion of the protein domains. No ‘breathing-mode’-like dynamics is observed. The presence of divalent ions, such as Ca-ions affect the dynamics of especially solvent exposed parts of the protein, but leave the asymmetric domain motion unaffected. Comparative study of the structural features and protein dynamics within membrane environment has been conducted on an already experimentally established structure of M2 protein from Influenza A virus. It is a single TMD bitopic tetrameric protein with 97-amino acid residues and with N-terminus directed toward the viral exterior. It displays proton conductance and is activated by low pH environment as found in endosomes. The principal architecture of proton conductance in M2 channels is believed to lie within its TMDs, in the form of pore-lining conserved HXXXW motif. The decrease in the pH is mimicked by increasing the protonation state of the bundle in order to reenact the activation state. At lower pH, ions are found to enter through the C-terminal and travel to the vicinity of pore-lining conserved HXXXW motif. The fluctuation of the histidine ring system is found to increase at lower pH while it has hardly any effect on the overall protein dynamics.