Broad-Spectrum Allosteric Inhibition of Herpesvirus Proteases

Gable, Jonathan E.; Lee, Gregory M.; Jaishankar, Priyadarshini; Hearn, Brian R.; Waddling, C.A.; Renslo, Adam R.; Craik, Charles S.

doi:10.1021/bi5003234

Cited by 15 publications

(28 citation statements)

References 50 publications

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“…This was predicted earlier and verified by our monomeric structure [75,80,82]. Additionally, there are some models of monomer-like structures of C-terminally truncated KSHVassemblin (D222-256) with inhibitors that bind to the dimerization area [81,84], see the discussion in the Inhibitors section below.…”

Section: Monomeric Structuresupporting

confidence: 84%

“…Dimerization induces refolding of a loop (residues 162 to 171) near the active site. In its conformation in dimeric assemblin, this loop stabilizes the oxyanion hole [75,[80][81][82]. This mechanism is essential to activate the assemblin.…”

Section: Regulation Of Activitymentioning

confidence: 99%

“…Kinetic IC 50 values of the best inhibitor (DD2) and some derivatives (Table 5) with assemblins from KSHV, EBV, HCMV and HSV-2 were compared [81]. The assemblins from KSHV, EBV (gammaherpesviruses) and HCMV (betaherpesvirus) were inhibited at similar rates with IC 50 values for the most potent inhibitor DD2 of 1.5, 7.7 and 7.4 µM, respectively.…”

Section: Inhibitorsmentioning

confidence: 99%

“…This might be due to lower affinity of the inhibitor or due to differences in the experimental conditions, because of weaker dimer stability in line with lower proteolytic activity of HSV-2-assemblin. Consequently, a different experimental setup had to be used for HSV-2 [81].…”

Section: Inhibitorsmentioning

confidence: 99%

“…Crystal structures of truncated KSHV-assemblin (D222-256) that was inhibited with DD2 or DD2 derivatives were determined [81,84]. In this KSHV-assemblin construct, the two C-terminal helices were removed by C-terminally truncating the assemblin.…”

Section: Inhibitorsmentioning

confidence: 99%

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Assemblins as maturational proteases in herpesviruses

Zühlsdorf

Hinrichs

2017

Journal of General Virology

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During assembly of herpesvirus capsids, a protein scaffold self-assembles to ring-like structures forming the scaffold of the spherical procapsids. Proteolytic activity of the herpesvirus maturational protease causes structural changes that result in angularization of the capsids. In those mature icosahedral capsids, the packaging of viral DNA into the capsids can take place. The strictly regulated protease is called assemblin. It is inactive in its monomeric state and activated by dimerization. The structures of the dimeric forms of several assemblins from all herpesvirus subfamilies have been elucidated in the last two decades. They revealed a unique serine-protease fold with a catalytic triad consisting of a serine and two histidines. Inhibitors that disturb dimerization by binding to the dimerization area were found recently. Additionally, the structure of the monomeric form of assemblin from pseudorabies virus and some monomer-like structures of Kaposi's sarcoma-associated herpesvirus assemblin were solved. These findings are the proof-of-principle for the development of new anti-herpesvirus drugs. Therefore, the most important information on this fascinating and unique class of proteases is summarized here.

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Section: Monomeric Structuresupporting

confidence: 84%

Section: Regulation Of Activitymentioning

confidence: 99%

Section: Inhibitorsmentioning

confidence: 99%

Section: Inhibitorsmentioning

confidence: 99%

Section: Inhibitorsmentioning

confidence: 99%

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Assemblins as maturational proteases in herpesviruses

Zühlsdorf

Hinrichs

2017

Journal of General Virology

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Fragment‐Based Protein–Protein Interaction Antagonists of a Viral Dimeric Protease

et al. 2016

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Fragment-based drug discovery has shown promise as an approach for challenging targets such as protein-protein interfaces. We developed and applied an activity-based fragment screen against dimeric Kaposi’s sarcoma-associated herpesvirus protease (KSHV Pr) using an optimized fluorogenic substrate. Dose response determination was performed as a confirmation screen and NMR spectroscopy was used to map fragment inhibitor binding to KSHV Pr. Kinetic assays demonstrated that several initial hits also inhibit human cytomegalovirus protease (HCMV Pr). Binding of these hits to HCMV Pr was also confirmed via NMR spectroscopy. Despite the use of a target-agnostic fragment library, more than 80% of confirmed hits disrupted dimerization and bound to a previously reported pocket at the dimer interface of KSHV Pr, not to the active site. One class of fragments, an aminothiazole scaffold, was further explored using commercially available analogs. These compounds demonstrated greater than 100-fold improvement of inhibition. This study illustrates the power of fragment-based screening for these challenging enzymatic targets and provides an example of the potential druggability of pockets at protein-protein interfaces.

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The effect of dimerization and ligand binding on the dynamics of Kaposi's sarcoma‐associated herpesvirus protease

Bern

Tobi

2022

Proteins

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The Kaposi's sarcoma-associated herpesvirus protease is essential for virus maturation. This protease functions under allosteric regulation that establishes its enzymatic activity upon dimerization. It exists in equilibrium between an inactive monomeric state and an active, weakly associating, dimeric state that is stabilized upon ligand binding. The dynamics of the protease dimer and its monomer were studied using the Gaussian network model and the anisotropic network model , and its role in mediating the allosteric regulation is demonstrated. We show that the dimer is composed of five dynamical domains. The central domain is formed upon dimerization and composed of helix five of each monomer, in addition to proximal and distal domains of each monomer. Dimerization reduces the mobility of the central domains and increases the mobility of the distal domains, in particular the binding site within them. The three slowest ANM modes of the dimer assist the protease in ligand binding, motion of the conserved Arg142 and Arg143 toward the oxyanion, and reducing the activation barrier for the tetrahedral transition state by stretching the bond that is cleaved by the protease. In addition, we show that ligand binding reduces the motion of helices α1 and α5 at the interface and explain how ligand binding can stabilize the dimer.

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Broad-Spectrum Allosteric Inhibition of Herpesvirus Proteases

Cited by 15 publications

References 50 publications

Assemblins as maturational proteases in herpesviruses

Assemblins as maturational proteases in herpesviruses

Fragment‐Based Protein–Protein Interaction Antagonists of a Viral Dimeric Protease

The effect of dimerization and ligand binding on the dynamics of Kaposi's sarcoma‐associated herpesvirus protease

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