Long-Range Communication between Different Functional Sites in the Picornaviral 3C Protein

Chan, Yan Mei; Moustafa, Ibrahim M.; Arnold, Jamie J.; Cameron, Craig Ε.; Boehr, David D.

doi:10.1016/j.str.2016.02.019

Cited by 10 publications

(11 citation statements)

References 58 publications

(82 reference statements)

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Figure 6 A shows the scaled scattering profiles of 3C at concentrations ranging from 3.2 to 13.0 mg/mL. The overlap of the scaled curves at the low ends of scattering angles was consistent with the protein remaining monomeric over this concentration range in agreement with the NMR and dynamic light scattering studies ( Chan et al., 2016 ). The scattering data at the low-angle end showed a linear correlation, satisfying the Guinier approximation (qR g < 1.3), from which we determined a radius of gyration (R g ) of 17.26 Å ( Figure 6 A).…”

Section: Resultssupporting

confidence: 77%

The RNA-Binding Site of Poliovirus 3C Protein Doubles as a Phosphoinositide-Binding Domain

et al. 2017

View full text Add to dashboard Cite

SUMMARY Some viruses use phosphatidylinositol phosphate (PIP) to mark membranes used for genome replication or virion assembly. PIP-binding motifs of cellular proteins do not exist in viral proteins. Molecular-docking simulations revealed a putative site of PIP binding to poliovirus (PV) 3C protein that was validated using NMR spectroscopy. The PIP-binding site was located on a highly dynamic α-helix that also functions in RNA binding. Broad PIP-binding activity was observed in solution using a fluorescence polarization assay or in the context of a lipid bilayer using an on-chip, fluorescence assay. All-atom molecular dynamics simulations of the 3C protein-membrane interface revealed PIP clustering and perhaps PIP-dependent conformations. PIP clustering was mediated by interaction with residues that interact with the RNA phosphodiester backbone. We conclude that 3C binding to membranes will be determined by PIP abundance. We suggest that the duality of function observed for 3C may extend to RNA-binding proteins of other viruses.

show abstract

Section: Resultssupporting

confidence: 77%

The RNA-Binding Site of Poliovirus 3C Protein Doubles as a Phosphoinositide-Binding Domain

et al. 2017

View full text Add to dashboard Cite

show abstract

“…Ile94 3C*D and Ile103 3C*D also do not appear to undergo detectable exchange, although the resonances are of low intensity and the relaxation dispersion profiles were noisy, likely due to the slow tumbling time, and hence elevated transverse relaxation rate, of 3CD. Ile94 3C is adjacent to the RNA and lipid binding site in 3C [ 20 , 21 , 91 ]. The presence of the linked 3D domain may result in conformational dynamic differences important for the RNA binding differences between 3C and 3CD [ 14 , 15 , 16 ].…”

Section: Discussionmentioning

confidence: 99%

The Picornavirus Precursor 3CD Has Different Conformational Dynamics Compared to 3Cpro and 3Dpol in Functionally Relevant Regions

Winston

Boehr

2021

Viruses

View full text Add to dashboard Cite

Viruses have evolved numerous strategies to maximize the use of their limited genetic material, including proteolytic cleavage of polyproteins to yield products with different functions. The poliovirus polyprotein 3CD is involved in important protein-protein, protein-RNA and protein-lipid interactions in viral replication and infection. It is a precursor to the 3C protease and 3D RNA-dependent RNA polymerase, but has different protease specificity, is not an active polymerase, and participates in other interactions differently than its processed products. These functional differences are poorly explained by the known X-ray crystal structures. It has been proposed that functional differences might be due to differences in conformational dynamics between 3C, 3D and 3CD. To address this possibility, we conducted nuclear magnetic resonance spectroscopy experiments, including multiple quantum relaxation dispersion, chemical exchange saturation transfer and methyl spin-spin relaxation, to probe conformational dynamics across multiple timescales. Indeed, these studies identified differences in conformational dynamics in functionally important regions, including enzyme active sites, and RNA and lipid binding sites. Expansion of the conformational ensemble available to 3CD may allow it to perform additional functions not observed in 3C and 3D alone despite having nearly identical lowest-energy structures.

show abstract

“…S4 B and C) indicates that this region was likely locked into an "rigidified inactive" state by YDF binding, which may account for the "activity loss" of HRV14 3C in the presence of YDF. How the RMSF of residues 63 to 67 and 93 to 96 are affected by binding of YDF is not clear as they are distant to either the YDF interface or the substrate cleft, but long-range perturbation between noncontact regions on 3C has been observed before by NMR experiments (14,28).…”

Section: Resultsmentioning

confidence: 99%

Inhibitory antibodies identify unique sites of therapeutic vulnerability in rhinovirus and other enteroviruses

Meng

Lan

Xie

et al. 2020

Proc. Natl. Acad. Sci. U.S.A.

View full text Add to dashboard Cite

The existence of multiple serotypes renders vaccine development challenging for most viruses in theEnterovirusgenus. An alternative and potentially more viable strategy for control of these viruses is to develop broad-spectrum antivirals by targeting highly conserved proteins that are indispensable for the virus life cycle, such as the 3C protease. Previously, two single-chain antibody fragments, YDF and GGVV, were reported to effectively inhibit human rhinovirus 14 proliferation. Here, we found that both single-chain antibody fragments target sites on the 3C protease that are distinct from its known drug site (peptidase active site) and possess different mechanisms of inhibition. YDF does not block the active site but instead noncompetitively inhibits 3C peptidase activity through an allosteric effect that is rarely seen for antibody protease inhibitors. Meanwhile, GGVV antagonizes the less-explored regulatory function of 3C in genome replication. The interaction between 3C and the viral genome 5′ noncoding region has been reported to be important for enterovirus genome replication. Here, the interface between human rhinovirus 14 3C and its 5′ noncoding region was probed by hydrogen–deuterium exchange coupled mass spectrometry and found to partially overlap with the interface between GGVV and 3C. Consistently, prebinding of GGVV completely abolishes interaction between human rhinovirus 14 3C and its 5′ noncoding region. The epitopes of YDF and GGVV, therefore, represent two additional sites of therapeutic vulnerability in rhinovirus. Importantly, the GGVV epitope appears to be conserved across many enteroviruses, suggesting that it is a promising target for pan-enterovirus inhibitor screening and design.

show abstract

Long-Range Communication between Different Functional Sites in the Picornaviral 3C Protein

Cited by 10 publications

References 58 publications

The RNA-Binding Site of Poliovirus 3C Protein Doubles as a Phosphoinositide-Binding Domain

The RNA-Binding Site of Poliovirus 3C Protein Doubles as a Phosphoinositide-Binding Domain

The Picornavirus Precursor 3CD Has Different Conformational Dynamics Compared to 3Cpro and 3Dpol in Functionally Relevant Regions

Inhibitory antibodies identify unique sites of therapeutic vulnerability in rhinovirus and other enteroviruses

Contact Info

Product

Resources

About