Tracking Higher Order Protein Structure by Hydrogen-Deuterium Exchange Mass Spectrometry

Benhaim, Mark A.; Lee, Kelly K.; Guttman, Miklós

doi:10.2174/0929866526666181212165037

Cited by 25 publications

(29 citation statements)

References 124 publications

(125 reference statements)

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“…HDX-MS is a solution state biophysical and structural technique that monitors the accessibility of amide hydrogens along the protein backbone. HDX-MS directly monitors dynamic structural changes and motion throughout a protein that are otherwise invisible to other structural approaches [45,50,82,83]. At low pH conditions approaching fusion activation, dynamic changes across the HA were observed where the HA1-HA1 trimeric interface became bolstered and the HA2 fusion peptide proximal subdomain became more dynamic.…”

Section: Despite Their Common Architectures Activation Mechanisms Anmentioning

confidence: 99%

New Biophysical Approaches Reveal the Dynamics and Mechanics of Type I Viral Fusion Machinery and Their Interplay with Membranes

Benhaim

Lee

2020

Viruses

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Protein-mediated membrane fusion is a highly regulated biological process essential for cellular and organismal functions and infection by enveloped viruses. During viral entry the membrane fusion reaction is catalyzed by specialized protein machinery on the viral surface. These viral fusion proteins undergo a series of dramatic structural changes during membrane fusion where they engage, remodel, and ultimately fuse with the host membrane. The structural and dynamic nature of these conformational changes and their impact on the membranes have long-eluded characterization. Recent advances in structural and biophysical methodologies have enabled researchers to directly observe viral fusion proteins as they carry out their functions during membrane fusion. Here we review the structure and function of type I viral fusion proteins and mechanisms of protein-mediated membrane fusion. We highlight how recent technological advances and new biophysical approaches are providing unprecedented new insight into the membrane fusion reaction.Viruses 2020, 12, 413 2 of 21 built. Type II fusion proteins are found in viruses including flaviviruses and alphaviruses such as Dengue, Zika, and Chikungunya viruses, and even have been identified in eukaryotic cell-cell fusion systems [9][10][11][12]. Type III fusion proteins are found in rhabdoviruses (such as Rabies and Vesicular Stomatatis virus G glycoproteins), herpesviruses (Herpes Simplex virus 1 gB protein), as well as baculovirus [12][13][14][15][16]. While the individual folds exhibited by these classes are completely different, they share common functional traits in that all adopt a pre-fusion conformation prior to activation in which one terminus of the protein is anchored in the virus membrane by a transmembrane domain and a second membrane active component, either a fusion peptide or loop, is sequestered from interacting with membranes ( Figure 1) [12]. A trigger or set of triggers, such as exposure to low pH in endosomes or receptor binding, spurs the machinery to reorganize into a post-fusion state in which the two membrane active components are colocalized.Until recently, static structures of the pre-and post-fusion states of isolated fusion protein ectodomains and biochemical or spectroscopic measurements were the primary pieces of information that informed our models of membrane fusion. While the static structures provide defined endpoints for the conformational change that drives membrane fusion, they do not tell us how these conformational changes occur or how these proteins interact with and perturb the lipid membrane during fusion. Likewise, fluorescence spectroscopy and circular dichroism studies have shown that HA-fusion activation leads to population of discernable intermediates rather than transitioning directly and irreversibly from pre-to post-fusion states [17][18][19][20]. These studies show that not all HAs respond to activation in the same way and some require different pH conditions to fully activate [20]. While such studies provided valuable information...

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Section: Despite Their Common Architectures Activation Mechanisms Anmentioning

confidence: 99%

New Biophysical Approaches Reveal the Dynamics and Mechanics of Type I Viral Fusion Machinery and Their Interplay with Membranes

Benhaim

Lee

2020

Viruses

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“…When ionized in a mass spectrometer different protein conformations take on charge differently due to their size and shape, thus allowing one to isolate and analyze target conformers and their peptide fragments . ECD also enables fragmentation of a peptide without scrambling of the deuterium label, as is the case with other fragmentation methods, meaning deuterium uptake can be analyzed with single amino acid resolution . Pan et al were able to take advantage of different conformers’ preferences for different charge states and select for specific isotopically labeled Aβ42 conformers using precursor ion selection enabling analysis of the Aβ42 oligomer's HDX profile with single amino acid resolution using ECD …”

Section: Monitoring Transiently Preferred Conformational States In Inmentioning

confidence: 99%

“…These motions range from small fluctuations in local structure to large‐scale conformational rearrangements between distinct structural states. Despite the fact that these dynamic motions and structural rearrangements are critical for protein function, we do not yet fully understand how these motions and conformational changes contribute to protein structure and function . Furthermore, high‐resolution models are usually only available for the endpoints of dynamic processes and one can only infer what takes place during protein motion and conformational changes.…”

Section: Introductionmentioning

confidence: 99%

“…Despite the fact that these dynamic motions and structural rearrangements are critical for protein function, we do not yet fully understand how these motions and conformational changes contribute to protein structure and function. 1 Furthermore, high-resolution models are usually only available for the endpoints of dynamic processes and one can only infer what takes place during protein motion and conformational changes. In order to fully understand how a protein functions, we need to understand the dynamic structural changes that lead to functionally distinct conformations.…”

Section: Introductionmentioning

confidence: 99%

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

2019

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Much of our understanding of protein structure and mechanistic function has been derived from static high‐resolution structures. As structural biology has continued to evolve it has become clear that high‐resolution structures alone are unable to fully capture the mechanistic basis for protein structure and function in solution. Recently Hydrogen/Deuterium‐exchange Mass Spectrometry (HDX‐MS) has developed into a powerful and versatile tool for structural biologists that provides novel insights into protein structure and function. HDX‐MS enables direct monitoring of a protein's structural fluctuations and conformational changes under native conditions in solution even as it is carrying out its functions. In this review, we focus on the use of HDX‐MS to monitor these dynamic changes in proteins. We examine how HDX‐MS has been applied to study protein structure and function in systems ranging from large, complex assemblies to intrinsically disordered proteins, and we discuss its use in probing conformational changes during protein folding and catalytic function. Statement for a Broad Audience The biophysical and structural characterization of proteins provides novel insight into their functionalities. Protein motions, ranging from small scale local fluctuations to larger concerted structural rearrangements, often determine protein function. Hydrogen/Deuterium‐exchange Mass Spectrometry (HDX‐MS) has proven a powerful biophysical tool capable of probing changes in protein structure and dynamic protein motions that are often invisible to most other techniques.

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“…GPCRs are involved in cell signal transduction and is an important target molecule in the drug development industry. HDX-MS provides valuable insight into the conformation of the GPCRs when bound to different ligands [ 63 ]. For example, one of the pioneering studies which applied HDX-MS to a member of the GPCR protein family took place in 2010, where Zhang et al studied detergent solubilized β2-Adrenergic GPCR (β2AR) alone and bound to the inverse agonist carazolol [ 64 ].…”

Section: Applications Of Hdx-msmentioning

confidence: 99%

HDX-MS: An Analytical Tool to Capture Protein Motion in Action

2020

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Virtually all protein functions in the cell, including pathogenic processes, require coordinated motion of atoms or domains, i.e., conformational dynamics. Understanding protein dynamics is therefore critical both for drug development and to learn about the underlying molecular causes of many diseases. Hydrogen–Deuterium Exchange Mass Spectrometry (HDX-MS) provides valuable information about protein dynamics, which is highly complementary to the static picture provided by conventional high-resolution structural tools (i.e., X-ray crystallography and structural NMR). The amount of protein required to carry out HDX-MS experiments is a fraction of the amount required by alternative biophysical techniques, which are also usually lower resolution. Use of HDX-MS is growing quickly both in industry and academia, and it has been successfully used in numerous drug and vaccine development efforts, with important roles in understanding allosteric effects and mapping binding sites.

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Tracking Higher Order Protein Structure by Hydrogen-Deuterium Exchange Mass Spectrometry

Cited by 25 publications

References 124 publications

New Biophysical Approaches Reveal the Dynamics and Mechanics of Type I Viral Fusion Machinery and Their Interplay with Membranes

New Biophysical Approaches Reveal the Dynamics and Mechanics of Type I Viral Fusion Machinery and Their Interplay with Membranes

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

HDX-MS: An Analytical Tool to Capture Protein Motion in Action

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