Hsu HJ and Fischer WB (2012), In silico investigations of possible routes of ...

XB-ART-43195

J Mol Model 2012 Feb 01;182:501-14. doi: 10.1007/s00894-011-1092-6.

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In silico investigations of possible routes of assembly of ORF 3a from SARS-CoV.

Hsu HJ , Fischer WB .

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ORF 3a of human severe acute respiratory syndrome corona virus (SARS-CoV) has been identified as a 274 amino acid membrane protein. When expressed in Xenopus oocytes the protein forms channels. Based on bioinformatics approaches the topology has been identified to include three transmembrane domains (TMDs). Since structural models from experiments are still lacking, computational methods can be challenged to generate such models. In this study, a 'sequential approach' for the assembly is proposed in which the individual TMDs are assembled one by one. This protocol is compared with a concerted protocol in which all TMDs are assembled simultaneously. The role of the loops between the TMDs during assembly of the monomers into a bundle is investigated. Molecular dynamics simulations for 20 ns are performed as a short equilibration to assess the bundle stability in a lipid environment. The results suggest that bundles are likely with the second TMD facing the putative pore. All the putative bundles show water molecules trapped within the lumen of the pore with only occasional events of complete crossing.

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Species referenced: Xenopus
Genes referenced: ttn

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References [+] :

Agirre, Viroporin-mediated membrane permeabilization. Pore formation by nonstructural poliovirus 2B protein. 2002, Pubmed

	Fig. 1. Flowchart of the steps involved forming the tetrameric assemblies. Single TMDs are either assembled in a sequential (Seq1, Seq2) or concerted (Sim) manner to form monomeric structures. The monomers are assembled into tetramers (T) either with loops (L) or without prior to assembly
	Fig. 2. Root mean square deviation (RMSD) of the Cα backbones of the single TMDs, TMD1, TMD2 and TMD3, referring to the respective starting structure (a). Root mean square fluctuation (RMSF) of the atoms of the amino acids (b). The TMDs are overlaid so that the atom numbers match for each TMD. Values for TMD1 are shown in light gray, those for TMD2 in gray and the values for TMD3 in black
	Fig. 3. Monomers according to the assembly protocol: Seq1 (a), Seq2 (b) and Sim (c). Hydrophilic residues are highlighted in blue, hydrophobic residues in green. All models are drawn in a ‘Gaussian Contact’ illustration (MOE)
	Fig. 4. Tetramers according to the assembly protocols without loops: T-Seq1 (a), T-Seq2 (b), T-Sim (c); tetramers assembled with loops added after monomer assembly: T-Seq1-L (d), T-Seq2-L (e), T-Sim-L (f)
	Fig. 5. Root mean square deviation (RMSD) of the of Cα backbones of the bundle structures referring to the starting structure. T-Seq1 (gray), T-Seq2 (light gray) and T-Sim are shown (aI). The respective RMSD values for the individual TMDs of each simulation (TMD1 in light gray, TMD2 in gray, TMD3 in black) are shown separately (aII-IV). RMSD values of the bundles including the loops are shown for T-Seq1-L, TSeq2-L and T-Sim-L (b). Color coding and arrangement of the panels like in (a)
	Fig. 6. Models of T-Seq1 (a), T-Seq2 (b) and T-Sim (c), T-Seq1-L (d), T-Seq2-L (e) and T-Sim-L (f) are shown in their starting conformation (green) and after 20 ns of MD simulation (red)
	Fig. 7. Pore radii calculated using the software HOLE [56]. The values of the first 25 structures, covering 500 ps simulation in steps of 20 ps, are averaged and depicted in light lines. A similar average has been calculated covering the last 500 ps of the simulations (thick lines). Models of T-Seq1 (a), T-Seq2 (b) and T-Sim (c), T-Seq1-L (d), T-Seq2-L (e) and T-Sim-L (f) are shown