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

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.

Bridging protein structure, dynamics, and function using hydrogen/deuterium‐exchange mass spectrometry

“…As the field continues to evolve, the technique is becoming increasingly powerful and accessible, further expanding the scale and complexity of systems capable of being studied. For example, HDX‐MS has recently been used to study the structural dynamics of proteins on intact enveloped viruses such as dengue virus and influenza virus . Similarly, the structural characterization of integral membrane proteins in native membrane environments by HDX‐MS has become commonplace.…”

“…For example, HDX-MS has recently been used to study the structural dynamics of proteins on intact enveloped viruses such as dengue virus and influenza virus. [61][62][63] Similarly, the structural characterization of integral membrane proteins in native membrane environments by HDX-MS has become commonplace. What were once considered pipedreams of structural biology are now within reach.…”

Plasma membrane association facilitates conformational changes in the Marburg virus protein VP40 dimer

“…32 By determining the solvent accessibility of the mVP40 residues, important residues involved in binding to the membrane, as well as those residues at the oligomerization interface were identified. However, the structural details of the mVP40 conformation after membrane association is still lacking.…”

The unmasking of the lipid binding face of sphingosine kinase 1

“…1). It seems likely this interface may protrude into the hydrophobic region of the plasma by guest, on May 11, 2018 www.jlr.org membrane or interact at the neck of budding endocytic vesicles where there may be some packing defects in the membrane. These hydrophobic interactions may contribute to the necessary site specific localization of SK1 at the plasma membrane complementing the interaction of Lys 27 , Lys 29 , and Arg 186 with PA.…”

“…In the absence of structural data with the lipid ligand bound, HDX-MS has proved to be a valuable method for dissecting the lipid binding interface of several peripheral proteins (11,12). HDX-MS is used to measure the rate of exchange of amide hydrogens with solvent in order to study protein structural dynamics.…”