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Detection of lipid-induced structural changes of the Marburg virus matrix protein VP40 using hydrogen/deuterium exchange-mass spectrometry

View Article: PubMed Central - PubMed

ABSTRACT

Marburg virus (MARV) is a lipid-enveloped virus from the Filoviridae family containing a negative sense RNA genome. One of the seven MARV genes encodes the matrix protein VP40, which forms a matrix layer beneath the plasma membrane inner leaflet to facilitate budding from the host cell. MARV VP40 (mVP40) has been shown to be a dimeric peripheral protein with a broad and flat basic surface that can associate with anionic phospholipids such as phosphatidylserine. Although a number of mVP40 cationic residues have been shown to facilitate binding to membranes containing anionic lipids, much less is known on how mVP40 assembles to form the matrix layer following membrane binding. Here we have used hydrogen/deuterium exchange (HDX) mass spectrometry to determine the solvent accessibility of mVP40 residues in the absence and presence of phosphatidylserine and phosphatidylinositol 4,5-bisphosphate. HDX analysis demonstrates that two basic loops in the mVP40 C-terminal domain make important contributions to anionic membrane binding and also reveals a potential oligomerization interface in the C-terminal domain as well as a conserved oligomerization interface in the mVP40 N-terminal domain. Lipid binding assays confirm the role of the two basic patches elucidated with HD/X measurements, whereas molecular dynamics simulations and membrane insertion measurements complement these studies to demonstrate that mVP40 does not appreciably insert into the hydrocarbon region of anionic membranes in contrast to the matrix protein from Ebola virus. Taken together, we propose a model by which association of the mVP40 dimer with the anionic plasma membrane facilitates assembly of mVP40 oligomers.

No MeSH data available.


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Hydrogen/deuterium exchange of mVP40 in the presence of PS containing vesicles.A, ribbon map indicating the percentage of hydrogen/deuterium exchange that occurred in peptide fragments over the entire exchange period. Each row corresponds to each time point from 10 to 100,000 s. Color coding indicates the percentage of deuterium incorporation in the given time point. B, percent labeling of the 10-s time point mapped onto the crystal structure of mVP40 (PDB code 5B0V). Color coding is as indicated in A. C, space filling models of mVP40 protein (right) and mVP40 after incubation with lipids. Side view of a mVP40 monomer is also shown for both conditions.
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Figure 7: Hydrogen/deuterium exchange of mVP40 in the presence of PS containing vesicles.A, ribbon map indicating the percentage of hydrogen/deuterium exchange that occurred in peptide fragments over the entire exchange period. Each row corresponds to each time point from 10 to 100,000 s. Color coding indicates the percentage of deuterium incorporation in the given time point. B, percent labeling of the 10-s time point mapped onto the crystal structure of mVP40 (PDB code 5B0V). Color coding is as indicated in A. C, space filling models of mVP40 protein (right) and mVP40 after incubation with lipids. Side view of a mVP40 monomer is also shown for both conditions.

Mentions: Some of the deuterium accumulation plots of peptide fragments showed significant changes in the deuteration levels in the presence of phospholipid membranes compared with the protein alone sample (Fig. 6). These peptide fragments do not originate from the dimer interface or basic patch of mVP40 and hence we hypothesize that these peptides arise from regions of mVP40 that undergo structural changes triggered by mVP40 interaction with phospholipid membranes. For further analysis of these regions we produced a rainbow map for mVP40 in the presence of membranes representing the percentage of hydrogen/deuterium exchange for each overlapping peptide for the entire length of reaction (Fig. 7A).


Detection of lipid-induced structural changes of the Marburg virus matrix protein VP40 using hydrogen/deuterium exchange-mass spectrometry
Hydrogen/deuterium exchange of mVP40 in the presence of PS containing vesicles.A, ribbon map indicating the percentage of hydrogen/deuterium exchange that occurred in peptide fragments over the entire exchange period. Each row corresponds to each time point from 10 to 100,000 s. Color coding indicates the percentage of deuterium incorporation in the given time point. B, percent labeling of the 10-s time point mapped onto the crystal structure of mVP40 (PDB code 5B0V). Color coding is as indicated in A. C, space filling models of mVP40 protein (right) and mVP40 after incubation with lipids. Side view of a mVP40 monomer is also shown for both conditions.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC5391744&req=5

Figure 7: Hydrogen/deuterium exchange of mVP40 in the presence of PS containing vesicles.A, ribbon map indicating the percentage of hydrogen/deuterium exchange that occurred in peptide fragments over the entire exchange period. Each row corresponds to each time point from 10 to 100,000 s. Color coding indicates the percentage of deuterium incorporation in the given time point. B, percent labeling of the 10-s time point mapped onto the crystal structure of mVP40 (PDB code 5B0V). Color coding is as indicated in A. C, space filling models of mVP40 protein (right) and mVP40 after incubation with lipids. Side view of a mVP40 monomer is also shown for both conditions.
Mentions: Some of the deuterium accumulation plots of peptide fragments showed significant changes in the deuteration levels in the presence of phospholipid membranes compared with the protein alone sample (Fig. 6). These peptide fragments do not originate from the dimer interface or basic patch of mVP40 and hence we hypothesize that these peptides arise from regions of mVP40 that undergo structural changes triggered by mVP40 interaction with phospholipid membranes. For further analysis of these regions we produced a rainbow map for mVP40 in the presence of membranes representing the percentage of hydrogen/deuterium exchange for each overlapping peptide for the entire length of reaction (Fig. 7A).

View Article: PubMed Central - PubMed

ABSTRACT

Marburg virus (MARV) is a lipid-enveloped virus from the Filoviridae family containing a negative sense RNA genome. One of the seven MARV genes encodes the matrix protein VP40, which forms a matrix layer beneath the plasma membrane inner leaflet to facilitate budding from the host cell. MARV VP40 (mVP40) has been shown to be a dimeric peripheral protein with a broad and flat basic surface that can associate with anionic phospholipids such as phosphatidylserine. Although a number of mVP40 cationic residues have been shown to facilitate binding to membranes containing anionic lipids, much less is known on how mVP40 assembles to form the matrix layer following membrane binding. Here we have used hydrogen/deuterium exchange (HDX) mass spectrometry to determine the solvent accessibility of mVP40 residues in the absence and presence of phosphatidylserine and phosphatidylinositol 4,5-bisphosphate. HDX analysis demonstrates that two basic loops in the mVP40 C-terminal domain make important contributions to anionic membrane binding and also reveals a potential oligomerization interface in the C-terminal domain as well as a conserved oligomerization interface in the mVP40 N-terminal domain. Lipid binding assays confirm the role of the two basic patches elucidated with HD/X measurements, whereas molecular dynamics simulations and membrane insertion measurements complement these studies to demonstrate that mVP40 does not appreciably insert into the hydrocarbon region of anionic membranes in contrast to the matrix protein from Ebola virus. Taken together, we propose a model by which association of the mVP40 dimer with the anionic plasma membrane facilitates assembly of mVP40 oligomers.

No MeSH data available.


Related in: MedlinePlus