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Title: The influence of environment : simulations of the influenza BM2 proton channel
Author: Rouse, Sarah L.
ISNI:       0000 0004 2724 1787
Awarding Body: University of Oxford
Current Institution: University of Oxford
Date of Award: 2012
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The influenza BM2 proton channel is a small, tetrameric α-helical membrane protein, which is vital in multiple stages of the life cycle of the virus. The transmembrane (TM) domain is responsible for modulating pH to facilitate release of viral genetic material whilst the cytoplasmic domain is purported to play a role in virus budding via interactions with another protein, BM1. Coarse-grained molecular dynamics (CGMD) simulations have been employed in several studies of α-helical membrane protein folding with success. This methodology is applied to the BM2 TM domain to generate a structure of the tetramer based on sequence alone. A converged tetrameric bundle with left-handed packing was generated and stable following conversion to atomistic representation and conventional atomistic MD simulation. Sequential protonation of His19 residues within the putative HxxxW gating motif led to subtle rearrangements in the packing of the helices, allowing formation of a water wire once three of the His19 residues were protonated. The influence of solubilising detergents on membrane protein structure is a topic of debate in the literature. The recent solution NMR structure of the BM2 TM domain was used as the basis of a study on the influence of two types of detergent micelle environment compared to a dipalmitoylphosphatidylcholine (DPPC) bilayer. The detergents chosen were zwitterionic dihexanoylphosphatidylcholine (DHPC) and non-ionic β-dodecylmaltoside (DDM). The results of these simulations suggest that BM2 is able to modulate its helix packing to compensate for hydrophobic mismatch in the DPPC bilayer. The protein-detergent complexes appeared to retain some of the features of detergent-only micelles. DDM detergent molecules replicated the binding mode of DPPC lipids (parallel to the protein bundle axis) whereas the DHPC detergent molecules adopted a binding mode perpendicular to the protein bundle axis. This appeared to influence the binding on/off rates of each detergent. Recent developments in the field of mass spectrometry have allowed intact membrane protein complexes in detergent micelles to be stabilised in the gas phase. Questions remain over the structure of these gas phase complexes compared to solution phase. A combination of non-equilibrium and equilibrium MD simulations was used to study the effects of transfer from solution to vacuum of protein-detergent complexes. The DHPC detergent micelle formed destabilising interactions with BM2 upon dehydration whereas the presence of the DDM micelle stabilised the solution phase protein conformation. In the final Chapter the multiscale simulation approach is used to study the interactions between BM2 and BM1. Protein-protein interactions formed via CGMD simulation were conserved upon atomistic simulation. The results of this thesis highlight the worth of multiscale simulation approaches in observing a greater range of timescales and processes.
Supervisor: Sansom, Mark S. P. Sponsor: Biotechnology and Biological Sciences Research Council
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available
Keywords: Computational biochemistry ; Membrane proteins ; Biochemistry ; Protein folding ; Biophysics ; Mass spectrometry ; Molecular biophysics (biochemistry) ; Biophysical chemistry