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Transmembrane domain of M2 protein from influenza A virus studied by solid-state (15)N polarization inversion spin
1National High Magnetic Field Laboratory, Florida State University, Tallahassee, Florida 32306-4005, USA.
Abstract:
The M2 protein from the influenza A virus forms a proton channel in the virion that is essential for infection. This tetrameric protein appears to form a four-helix bundle spanning the viral membrane. Here the solid-state NMR method, 2D polarization inversion spin exchange at magic angle (PISEMA), has been used to obtain multiple constraints from specifically amino acid-labeled samples. The improvement of spectral resolution from 2D PISEMA over 1D methods and 2D separated local field methods is substantial. The reliability of the method is validated by comparison of anisotropic chemical shift and heteronuclear dipolar interactions from single site labeled samples. The quantitative interpretation of the high-resolution constraints confirms the helix tilt to be within the range of previous experimental determinations (32 degrees -38 degrees ). The binding of the channel inhibitor, amantadine, results in no change in the backbone structure at position Val(27,28), which is thought to be a potential binding site for the inhibitor.
Insights
Solid-state NMR revealed the M2 proton channel structure in influenza A virus. Amantadine binding did not alter the backbone structure at a key site.
Area of Science:
- Structural biology
- Virology
- Biophysics
Background:
- The M2 protein of influenza A virus forms a proton channel crucial for viral infection.
- This tetrameric protein forms a four-helix bundle within the viral membrane.
Purpose of the Study:
- To determine the structural constraints of the M2 proton channel using solid-state NMR.
- To investigate the effect of amantadine binding on the M2 protein structure.
Main Methods:
- Utilized 2D polarization inversion spin exchange at magic angle (PISEMA), a solid-state NMR technique.
- Employed specifically amino acid-labeled samples to obtain multiple structural constraints.
- Validated method reliability through anisotropic chemical shift and heteronuclear dipolar interactions.
Main Results:
- Achieved substantial spectral resolution improvement compared to 1D and 2D separated local field methods.
- Quantitatively interpreted high-resolution constraints, confirming helix tilt between 32-38 degrees.
- Observed no change in backbone structure at Val(27,28) upon amantadine binding.
Conclusions:
- Solid-state NMR (PISEMA) provides high-resolution structural constraints for the influenza M2 proton channel.
- The M2 protein helix tilt is consistent with previous experimental findings.
- Amantadine does not appear to alter the M2 backbone structure at the investigated site.