Investigation of the Curvature Induction and Membrane Localization of the Influenza Virus M2 Protein Using Static and Off-Magic-Angle Spinning Solid-State Nuclear Magnetic Resonance of Oriented Bicelles
A wide variety of membrane proteins induce membrane curvature for function; thus, it is important to develop new methods to simultaneously determine membrane curvature and protein binding sites in membranes with multiple curvatures. We introduce solid-state nuclear magnetic resonance (NMR) methods b...
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| Other Authors: | |
| Format: | Article |
| Language: | en_US |
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American Chemical Society (ACS)
2016
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| Online Access: | http://hdl.handle.net/1721.1/105324 https://orcid.org/0000-0002-1801-924X https://orcid.org/0000-0001-5255-5858 |
| _version_ | 1811074925986840576 |
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| author | Wang, Tuo Hong, Mei |
| author2 | Massachusetts Institute of Technology. Department of Chemistry |
| author_facet | Massachusetts Institute of Technology. Department of Chemistry Wang, Tuo Hong, Mei |
| author_sort | Wang, Tuo |
| collection | MIT |
| description | A wide variety of membrane proteins induce membrane curvature for function; thus, it is important to develop new methods to simultaneously determine membrane curvature and protein binding sites in membranes with multiple curvatures. We introduce solid-state nuclear magnetic resonance (NMR) methods based on magnetically oriented bicelles and off-magic-angle spinning (OMAS) to measure membrane curvature and the binding site of proteins in mixed-curvature membranes. We demonstrate these methods on the influenza virus M2 protein, which not only acts as a proton channel but also mediates virus assembly and membrane scission. An M2 peptide encompassing the transmembrane (TM) domain and an amphipathic helix, M2(21–61), was studied and compared with the TM peptide (M2TM). Static [superscript 31]P NMR spectra of magnetically oriented 1,2-dimyristoyl-sn-glycero-3-phosphocholine (DMPC)/1,2-dihexanoyl-sn-glycero-3-phosphocholine (DHPC) bicelles exhibit a temperature-independent isotropic chemical shift in the presence of M2(21–61) but not M2TM, indicating that the amphipathic helix confers the ability to generate a high-curvature phase. Two-dimensional (2D) [superscript 31]P spectra indicate that this high-curvature phase is associated with the DHPC bicelle edges, suggestive of the structure of budding viruses from the host cell. [superscript 31]P- and [superscript 13]C-detected [superscript 1]H relaxation times of the lipids indicate that the majority of M2(21–61) is bound to the high-curvature phase. Using OMAS experiments, we resolved the [superscript 31]P signals of lipids with identical headgroups based on their distinct chemical shift anisotropies. On the basis of this resolution, 2D [superscript 1]H–[superscript 31]P correlation spectra show that the amide protons in M2(21–61) correlate with the DMPC but not DHPC [superscript 31]P signal of the bicelle, indicating that a small percentage of M2(21–61) partitions into the planar region of the bicelles. These results show that the amphipathic helix induces high membrane curvature and localizes the protein to this phase, in good agreement with the membrane scission function of the protein. These bicelle-based relaxation and OMAS solid-state NMR techniques are generally applicable to curvature-inducing membrane proteins such as those involved in membrane trafficking, membrane fusion, and cell division. |
| first_indexed | 2024-09-23T09:57:26Z |
| format | Article |
| id | mit-1721.1/105324 |
| institution | Massachusetts Institute of Technology |
| language | en_US |
| last_indexed | 2024-09-23T09:57:26Z |
| publishDate | 2016 |
| publisher | American Chemical Society (ACS) |
| record_format | dspace |
| spelling | mit-1721.1/1053242022-09-26T14:50:43Z Investigation of the Curvature Induction and Membrane Localization of the Influenza Virus M2 Protein Using Static and Off-Magic-Angle Spinning Solid-State Nuclear Magnetic Resonance of Oriented Bicelles Wang, Tuo Hong, Mei Massachusetts Institute of Technology. Department of Chemistry Wang, Tuo Hong, Mei A wide variety of membrane proteins induce membrane curvature for function; thus, it is important to develop new methods to simultaneously determine membrane curvature and protein binding sites in membranes with multiple curvatures. We introduce solid-state nuclear magnetic resonance (NMR) methods based on magnetically oriented bicelles and off-magic-angle spinning (OMAS) to measure membrane curvature and the binding site of proteins in mixed-curvature membranes. We demonstrate these methods on the influenza virus M2 protein, which not only acts as a proton channel but also mediates virus assembly and membrane scission. An M2 peptide encompassing the transmembrane (TM) domain and an amphipathic helix, M2(21–61), was studied and compared with the TM peptide (M2TM). Static [superscript 31]P NMR spectra of magnetically oriented 1,2-dimyristoyl-sn-glycero-3-phosphocholine (DMPC)/1,2-dihexanoyl-sn-glycero-3-phosphocholine (DHPC) bicelles exhibit a temperature-independent isotropic chemical shift in the presence of M2(21–61) but not M2TM, indicating that the amphipathic helix confers the ability to generate a high-curvature phase. Two-dimensional (2D) [superscript 31]P spectra indicate that this high-curvature phase is associated with the DHPC bicelle edges, suggestive of the structure of budding viruses from the host cell. [superscript 31]P- and [superscript 13]C-detected [superscript 1]H relaxation times of the lipids indicate that the majority of M2(21–61) is bound to the high-curvature phase. Using OMAS experiments, we resolved the [superscript 31]P signals of lipids with identical headgroups based on their distinct chemical shift anisotropies. On the basis of this resolution, 2D [superscript 1]H–[superscript 31]P correlation spectra show that the amide protons in M2(21–61) correlate with the DMPC but not DHPC [superscript 31]P signal of the bicelle, indicating that a small percentage of M2(21–61) partitions into the planar region of the bicelles. These results show that the amphipathic helix induces high membrane curvature and localizes the protein to this phase, in good agreement with the membrane scission function of the protein. These bicelle-based relaxation and OMAS solid-state NMR techniques are generally applicable to curvature-inducing membrane proteins such as those involved in membrane trafficking, membrane fusion, and cell division. National Institutes of Health (U.S.) (NIH Grant GM088204) 2016-11-14T20:01:42Z 2016-11-14T20:01:42Z 2015-03 2015-03 Article http://purl.org/eprint/type/JournalArticle 0006-2960 1520-4995 http://hdl.handle.net/1721.1/105324 Wang, Tuo, and Mei Hong. "Investigation of the Curvature Induction and Membrane Localization of the Influenza Virus M2 Protein Using Static and Off-Magic-Angle Spinning Solid-State Nuclear Magnetic Resonance of Oriented Bicelles." Biochemistry 54:13 (2015), pp. 2214-2226. https://orcid.org/0000-0002-1801-924X https://orcid.org/0000-0001-5255-5858 en_US http://dx.doi.org/10.1021/acs.biochem.5b00127 Biochemistry Article is made available in accordance with the publisher's policy and may be subject to US copyright law. Please refer to the publisher's site for terms of use. application/pdf American Chemical Society (ACS) PMC |
| spellingShingle | Wang, Tuo Hong, Mei Investigation of the Curvature Induction and Membrane Localization of the Influenza Virus M2 Protein Using Static and Off-Magic-Angle Spinning Solid-State Nuclear Magnetic Resonance of Oriented Bicelles |
| title | Investigation of the Curvature Induction and Membrane Localization of the Influenza Virus M2 Protein Using Static and Off-Magic-Angle Spinning Solid-State Nuclear Magnetic Resonance of Oriented Bicelles |
| title_full | Investigation of the Curvature Induction and Membrane Localization of the Influenza Virus M2 Protein Using Static and Off-Magic-Angle Spinning Solid-State Nuclear Magnetic Resonance of Oriented Bicelles |
| title_fullStr | Investigation of the Curvature Induction and Membrane Localization of the Influenza Virus M2 Protein Using Static and Off-Magic-Angle Spinning Solid-State Nuclear Magnetic Resonance of Oriented Bicelles |
| title_full_unstemmed | Investigation of the Curvature Induction and Membrane Localization of the Influenza Virus M2 Protein Using Static and Off-Magic-Angle Spinning Solid-State Nuclear Magnetic Resonance of Oriented Bicelles |
| title_short | Investigation of the Curvature Induction and Membrane Localization of the Influenza Virus M2 Protein Using Static and Off-Magic-Angle Spinning Solid-State Nuclear Magnetic Resonance of Oriented Bicelles |
| title_sort | investigation of the curvature induction and membrane localization of the influenza virus m2 protein using static and off magic angle spinning solid state nuclear magnetic resonance of oriented bicelles |
| url | http://hdl.handle.net/1721.1/105324 https://orcid.org/0000-0002-1801-924X https://orcid.org/0000-0001-5255-5858 |
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