Human antibody C10 neutralizes by diminishing Zika but enhancing dengue virus dynamics.

The human monoclonal antibody (HmAb) C10 potently cross-neutralizes Zika virus (ZIKV) and dengue virus. Analysis of antibody fragment (Fab) C10 interactions with ZIKV and dengue virus serotype 2 (DENV2) particles by cryoelectron microscopy (cryo-EM) and amide hydrogen/deuterium exchange mass spectrometry (HDXMS) shows that Fab C10 binding decreases overall ZIKV particle dynamics, whereas with DENV2, the same Fab causes increased dynamics. Testing of different Fab C10:DENV2 E protein molar ratios revealed that, at higher Fab ratios, especially at saturated concentrations, the Fab enhanced viral dynamics (detected by HDXMS), and observation under cryo-EM showed increased numbers of distorted particles.

Capsid protein structure in Zika virus reveals the flavivirus assembly process.

Structures of flavivirus (dengue virus and Zika virus) particles are known to near-atomic resolution and show detailed structure and arrangement of their surface proteins (E and prM in immature virus or M in mature virus). By contrast, the arrangement of the capsid proteins:RNA complex, which forms the core of the particle, is poorly understood, likely due to inherent dynamics. Here, we stabilize immature Zika virus via an antibody that binds across the E and prM proteins, resulting in a subnanometer resolution structure of capsid proteins within the virus particle.

Infectivity of Dengue Virus Serotypes 1 and 2 Is Correlated with E-Protein Intrinsic Dynamics but Not to Envelope Conformations.

Dengue is a mosquito-borne virus with dire health and economic impacts. Dengue is responsible for an estimated 390 million infections per year, with dengue 2 (DENV2) being the most virulent strain among the four serotypes. Interestingly, it is also in strains of this serotype that temperature-dependent large-scale morphological changes, termed "breathing," have been observed.

Uncovering metastability and disassembly hotspots in whole viral particles.

Viruses are metastable macromolecular assemblies that toggle between multiple conformational states through molecular rearrangements that are critical for mediating viral host entry. Viruses respond to different host specific environmental cues to form disassembly intermediates for the eventual release of genomic material required for replication. Although static snapshots of these intermediates have been captured through structural techniques such as X-ray crystallography and cryo-EM, the mechanistic details of these conformational rearrangements underpinning viral metastability have been poorly understood.

Mechanism of Enhanced Immature Dengue Virus Attachment to Endosomal Membrane Induced by prM Antibody.

Dengue virus (DENV) particles are released from cells in different maturation states. Fully immature DENV (immDENV) is generally non-infectious, but can become infectious when complexed with anti-precursor membrane (prM) protein antibodies. It is unknown how anti-prM antibody-coated particles can undergo membrane fusion since the prM caps the envelope (E) protein fusion loop.

Partial Intrinsic Disorder Governs the Dengue Capsid Protein Conformational Ensemble.

The 11 kDa, positively charged dengue capsid protein (C protein) exists stably as a homodimer and colocalizes with the viral genome within mature viral particles. Its core is composed of four alpha helices encompassing a small hydrophobic patch that may interact with lipids, but approximately 20% of the protein at the N-terminus is intrinsically disordered, making it challenging to elucidate its conformational landscape. Here, we combine small-angle X-ray scattering (SAXS), amide hydrogen-deuterium exchange mass spectrometry (HDXMS), and atomic-resolution molecular dynamics (MD) simulations to probe the dynamics of dengue C proteins.

Epitope and Paratope Mapping Reveals Temperature-Dependent Alterations in the Dengue-Antibody Interface.

Uncovering mechanisms of antibody-mediated neutralization for viral infections requires epitope and paratope mapping in the context of whole viral particle interactions with the antibody in solution. In this study, we use amide hydrogen/deuterium exchange mass spectrometry to describe the interface of a dengue virus-neutralizing antibody, 2D22, with its target epitope. 2D22 binds specifically to DENV2, a serotype showing strain-specific structural expansion at human host physiological temperatures of 37°C.

Conformational changes in intact dengue virus reveal serotype-specific expansion.

Dengue virus serotype 2 (DENV2) alone undergoes structural expansion at 37 °C (associated with host entry), despite high sequence and structural homology among the four known serotypes. The basis for this differential expansion across strains and serotypes is unknown and necessitates mapping of the dynamics of dengue whole viral particles to describe their coordinated motions and conformational changes when exposed to host-like environments. Here we capture the dynamics of intact viral particles of two serotypes, DENV1 and DENV2, by amide hydrogen/deuterium exchange mass spectrometry (HDXMS) and time resolved Förster Resonance Energy Transfer.

Active site coupling in PDE:PKA complexes promotes resetting of mammalian cAMP signaling.

Cyclic 3'5' adenosine monophosphate (cAMP)-dependent-protein kinase (PKA) signaling is a fundamental regulatory pathway for mediating cellular responses to hormonal stimuli. The pathway is activated by high-affinity association of cAMP with the regulatory subunit of PKA and signal termination is achieved upon cAMP dissociation from PKA. Although steps in the activation phase are well understood, little is known on how signal termination/resetting occurs.