Tripeptide Mimetics Inhibit the 20 S Proteasome by Covalent Bonding to the Active Threonines

Braun, Hannes A.; Umbreen, Sumaira; Groll, M.; Kuckelkorn, Ulrike; Młynarczuk, Izabela; Wigand, Moritz E.; Drung, Ilse; Kloetzel, Peter‐Michael; Schmidt, Boris

doi:10.1074/jbc.m502453200

Cited by 70 publications

(63 citation statements)

References 30 publications

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“…Samples were stored at −80° C until assayed for determination of proteasome peptidase activities. Samples were homogenized (1:10 w/v) in an ice-chilled buffer containing: 20 mM Tris-HCl (pH 7.2), 1 mM EDTA, 1mM sodium azide, 1 mM DTT, 0.1% NP40, and a 1:200 protease inhibitor cocktail (Sigma) (Braun et al, 2005). Homogenates were centrifuged at 14000g for 5 min at 4°C and the supernatants were used for determination of proteasome peptidase activity.…”

Section: Brain Proteasome Activitymentioning

confidence: 99%

Systemic administration of a proteasome inhibitor does not cause nigrostriatal dopamine degeneration

et al. 2007

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Proteasomal dysfunction has been suggested to contribute to the degeneration of nigrostriatal dopamine neurons in Parkinson's disease. A recent study reported that systemic treatment of rats with the proteasome inhibitor Z-lle-Glu(OtBu)-Ala-Leu-al (PSI) causes a slowly progressive degeneration of nigrostriatal dopamine neurons, the presence of inclusion bodies in dopamine neurons, and motor impairment. We examined in vitro and in vivo the effects of PSI on nigrostriatal dopamine neurons. Mass spectrometric analysis was employed to verify the authenticity of the PSI compound. PSI was non-specifically toxic to neurons in ventral mesencephalic organotypic slice cultures, indicating that impairment of proteasome function in vitro is toxic. Moreover, systemic administration of PSI transiently decreased brain proteasome activity. Systemic treatment of rats with PSI did not, however, result in any biochemical or anatomical evidence of lesions of nigrostriatal dopamine neurons, nor were any changes in locomotor activity observed. These data suggest that systemic administration of proteasome inhibitors to normal adult rats does not reliably cause an animal model of parkinsonism.

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Section: Brain Proteasome Activitymentioning

confidence: 99%

Systemic administration of a proteasome inhibitor does not cause nigrostriatal dopamine degeneration

et al. 2007

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“…Proteasome and PA28-20S proteasome was isolated from T2 cells (c20S), and PA28 was isolated from red blood (12,13).…”

Section: Methodsmentioning

confidence: 99%

The N-terminal Flanking Region of the TRP2360–368 Melanoma Antigen Determines Proteasome Activator PA28 Requirement for Epitope Liberation

Textoris-Taube

Henklein

Pollmann

et al. 2007

Journal of Biological Chemistry

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Proteasomes are known to produce major histocompatibility complex (MHC) class I ligands from endogenous antigens. The interferon-␥-inducible proteasome activator PA28 plays an important role in the generation of MHC ligands by proteasomes. Generation of the HLA-A*0201 restricted melanoma antigen TRP2 360 -368 by the proteasome has been shown to be dependent on the function of PA28 in vitro and in vivo (Sun, Y., Sijts, A. J., Song, M., Janek, K., Nussbaum, A. K., Kral, S., Schirle, M., Stevanovic, S., Paschen, A., Schild, H., Kloetzel, P. M., and Schadendorf, D. (2002) Cancer Res. 62, 2875-2882). Here we analyzed the role of the epitope sequence environment in determining this PA28 dependence. Experiments using the melanoma TRP2 288 -296 epitope and the murine cytomegalovirus-derived pp89 epitope precursor peptide for epitope replacement revealed that the TRP2 360 -368 flanking sequences can transfer PA28 dependence onto otherwise PA28 independent epitopes. Moreover, the N-terminal flanking sequence is sufficient to establish PA28 dependence of an epitope by allowing PA28-induced coordinated dual cleavages. These results show that N-terminal flanking sequences strongly influence epitope generation efficiency and that PA28 function is particularly relevant for the generation of normally poorly excised peptide products.The generation of major histocompatibility complex (MHC) 2 class I ligands and the recognition of MHC class I-ligand complexes by CD8ϩ T cells is an effective tool for the elimination of infected or disordered cells from organisms. The ubiquitin-proteasome system represents the major source for MHC class I-presented peptides exposed to CD8ϩ T cells (1). Ubiquitin-proteasome system-mediated peptide generation is influenced by structural changes in the 20S catalytic core (20S proteasome), which occur by exchange of the standard catalytic subunits by immunosubunits. The ATP-dependent 26S proteasome, composed of a 20S core and a 19S regulatory complex, recognizes substrates by a multi-ubiquitin signal. In contrast, 20S proteasomes can accept larger non-ubiquitinated peptides as substrates for ATP-independent degradation (3). However, 20S proteasomes exist in a latent state within the cells. In consequence, proteasome activity and function is regulated in part by modulating substrate entry by the PA28 complex, which attaches ATP-independent to the outer ␣-rings of the 20S proteasome. Detailed kinetic analysis using fluorogenic peptide substrates has shown that activation of the 20S proteasome by PA28 occurs by facilitating either substrate entry and/or product exit, with no effects on the active sites (3). In support, structural analyses have demonstrated that attachment of the activator to the 20S core leads to the opening of the central gate of the ␣-ring (4).The expression of PA28 is constitutively enhanced in cells with specialized antigen-presenting function and is induced by stimulation of cells with interferon-␥. The presentation of a number of viral MHC class I epitopes is enhanced in the presence...

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“…Protein content was estimated by BCA (Pierce, Rockford, IL). Proteasomal activity was measured as described previously (10). One hundred nanograms of purified proteasomes (13) were exposed to proteasome inhibitors and activity was determined with 100 Amol/L fluorogenic substrate [succinyl-Leu-Leu-Val-Tyr-7-amido-4-methylcoumarin (suc-LLVY-AMC), Bachem, Weil am Rhein, Germany].…”

Section: Methodsmentioning

confidence: 99%

“…In particular, we examined cellular effects of a new proteasome inhibitor, BSc2118 (10). Our data show that proteasome inhibitor treatment of cisplatin-sensitive and cisplatin-resistant melanoma cells induces cell death at concentrations that are not toxic to primary human fibroblasts.…”

Section: Introductionmentioning

confidence: 99%

Combined Effect of Proteasome and Calpain Inhibition on Cisplatin-Resistant Human Melanoma Cells

Młynarczuk-Biały

Roeckmann²,

Kuckelkorn

et al. 2006

Cancer Research

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Tripeptide Mimetics Inhibit the 20 S Proteasome by Covalent Bonding to the Active Threonines

Cited by 70 publications

References 30 publications

Systemic administration of a proteasome inhibitor does not cause nigrostriatal dopamine degeneration

Systemic administration of a proteasome inhibitor does not cause nigrostriatal dopamine degeneration

The N-terminal Flanking Region of the TRP2360–368 Melanoma Antigen Determines Proteasome Activator PA28 Requirement for Epitope Liberation

Combined Effect of Proteasome and Calpain Inhibition on Cisplatin-Resistant Human Melanoma Cells

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