N,C-Capped Dipeptides with Selectivity for Mycobacterial Proteasome over Human Proteasomes: Role of S3 and S1 Binding Pockets

Lin, Gang; Chidawanyika, Tamutenda; Tsu, Christopher; Warrier, Thulasi; Vaubourgeix, Julien; Blackburn, Christopher; Gigstad, Kenneth M.; Sintchak, Michael D.; Dick, Lawrence R.; Nathan, Carl

doi:10.1021/ja400021x

Cited by 52 publications

(92 citation statements)

References 18 publications

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“…1C), similar to DPLG-2 20 . Based on the dose-dependence curves, we calculated the IC 50 values of these compounds (Table 1) against Mtb20SOG; all were in the narrow range of 0.010–0.041 μM.…”

Section: Resultssupporting

confidence: 53%

“…IC 50 values of compounds against Mtb20SOG, β5i and β5c were determined from tests in a 96-well format as reported 20 . In brief, 1 μL of compound in a 3x series dilution in DMSO at concentrations from 100 μM to 0.0017 μM were spotted to the bottom of a black 96-well plate having a solid bottom.…”

Section: Methodsmentioning

confidence: 99%

“…In previous work, we screened 1600 N,C-capped dipeptides that varied in the P1 (the C-cap), P2, P3, and P4 (the N-cap) residues; variants in the P2, and P3 residues included both natural and unnatural amino acids. We identified several compounds that had 100-fold to over 1000-fold more selectivity for the Mtb 20S “open gate” CP (Mtb20SOG, in which the amino terminal heptapeptide is deleted from the αchains) than for the human c-20S β5 20 . One of the compounds, DPLG-2, was shown to penetrate M. tuberculosis to exert bactericidal activity against nonreplicating bacteria under nitrosative stress.…”

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confidence: 99%

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Structural Basis for the Species-Selective Binding of N,C-Capped Dipeptides to the Mycobacterium tuberculosis Proteasome

Hsu

Singh²,

Fan³

et al. 2016

Biochemistry

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The Mycobacterium tuberculosis (Mtb) 20S proteasome is vital for the pathogen to survive under nitrosative stress in vitro and to persist in mice. To qualify for drug development, inhibitors targeting Mtb 20S must spare both the human constitutive proteasome (c-20S) and immunoproteasome (i-20S). We recently reported members of a family of noncovalently binding dipeptide proteasome inhibitors that are highly potent and selective for Mtb 20S over human c-20S and i-20S. To understand the structural basis of their potency and selectivity, we have studied the structure–activity relationship of six derivatives and solved their co-crystal structures with Mtb 20S. The dipeptide inhibitors form an antiparallel β-strand with the active site β-strands. Selectivity is conferred by several features of Mtb 20S relative to its mouse counterparts, including a larger S1 pocket, additional hydrogen bonds in the S3 pocket, and hydrophobic interactions in the S4 pocket. Serine-20 and glutamine-22 of Mtb 20S interact with the dipeptides and confer Mtb-specific inhibition over c-20S and i-20S. The Mtb 20S and mammalian i-20S have a serine-27 that interacts strongly with the dipeptides, potentially explaining the higher inhibitory activity of the dipeptides toward i-20S over c-20S. This detailed structural knowledge will aid in optimizing the dipeptides as anti-tuberculosis drugs.

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Section: Resultssupporting

confidence: 53%

Section: Methodsmentioning

confidence: 99%

mentioning

confidence: 99%

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Structural Basis for the Species-Selective Binding of N,C-Capped Dipeptides to the Mycobacterium tuberculosis Proteasome

Hsu

Singh²,

Fan³

et al. 2016

Biochemistry

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“…The potential to target persistent or latent bacteria has made the Mtb proteasome system a prioritized target for the development of antituberculosis drugs (10,11). Indeed, Mtb-specific proteasome inhibitors have been identified that may provide a promising lead for new drugs to treat tuberculosis (12,13).…”

mentioning

confidence: 99%

Structural analysis of the dodecameric proteasome activator PafE in Mycobacterium tuberculosis

Bai

Wang

et al. 2016

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

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The human pathogen Mycobacterium tuberculosis (Mtb) requires a proteasome system to cause lethal infections in mice. We recently found that proteasome accessory factor E (PafE, Rv3780) activates proteolysis by the Mtb proteasome independently of adenosine triphosphate (ATP). Moreover, PafE contributes to the heat-shock response and virulence of Mtb. Here, we show that PafE subunits formed four-helix bundles similar to those of the eukaryotic ATP-independent proteasome activator subunits of PA26 and PA28. However, unlike any other known proteasome activator, PafE formed dodecamers with 12-fold symmetry, which required a glycine-XXX-glycine-XXX-glycine motif that is not found in previously described activators. Intriguingly, the truncation of the PafE carboxyl-terminus resulted in the robust binding of PafE rings to native proteasome core particles and substantially increased proteasomal activity, suggesting that the extended carboxyl-terminus of this cofactor confers suboptimal binding to the proteasome core particle. Collectively, our data show that proteasomal activation is not limited to hexameric ATPases in bacteria.

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“…The synthetic route and methods of compound characterization were similar to those reported for N,C-capped dipeptidomimetics (15,28), with modifications as given in the supplemental online material. All compounds were >95% pure.…”

Section: Methodsmentioning

confidence: 99%

Brief treatment with a highly selective immunoproteasome inhibitor promotes long-term cardiac allograft acceptance in mice

Karreci

Fan

Uehara

et al. 2016

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

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Constitutive proteasomes (c-20S) are ubiquitously expressed cellular proteases that degrade polyubiquitinated proteins and regulate cell functions. An isoform of proteasome, the immunoproteasome (i-20S), is highly expressed in human T cells, dendritic cells (DCs), and B cells, suggesting that it could be a potential target for inflammatory diseases, including those involving autoimmunity and alloimmunity. Here, we describe DPLG3, a rationally designed, noncovalent inhibitor of the immunoproteasome chymotryptic subunit β5i that has thousands-fold selectivity over constitutive β5c. DPLG3 suppressed cytokine release from blood mononuclear cells and the activation of DCs and T cells, diminished accumulation of effector T cells, promoted expression of exhaustion and coinhibitory markers on T cells, and synergized with CTLA4-Ig to promote long-term acceptance of cardiac allografts across a major histocompatibility barrier. These findings demonstrate the potential value of using brief posttransplant immunoproteasome inhibition to entrain a long-term response favorable to allograft survival as part of an immunomodulatory regimen that is neither broadly immunosuppressive nor toxic.

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N,C-Capped Dipeptides with Selectivity for Mycobacterial Proteasome over Human Proteasomes: Role of S3 and S1 Binding Pockets

Cited by 52 publications

References 18 publications

Structural Basis for the Species-Selective Binding of N,C-Capped Dipeptides to the Mycobacterium tuberculosis Proteasome

Structural Basis for the Species-Selective Binding of N,C-Capped Dipeptides to the Mycobacterium tuberculosis Proteasome

Structural analysis of the dodecameric proteasome activator PafE in Mycobacterium tuberculosis

Brief treatment with a highly selective immunoproteasome inhibitor promotes long-term cardiac allograft acceptance in mice

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