The Structure of the Mammalian 20S Proteasome at 2.75 Å Resolution

Abstract

The 20S proteasome is the catalytic portion of the 26S proteasome. Constitutively expressed mammalian 20S proteasomes have three active subunits, beta 1, beta 2, and beta 5, which are replaced in the immunoproteasome by interferon-gamma-inducible subunits beta 1i, beta 2i, and beta 5i, respectively. Here we determined the crystal structure of the bovine 20S proteasome at 2.75 A resolution. The structures of alpha 2, beta 1, beta 5, beta 6, and beta 7 subunits of the bovine enzyme were different from the yeast enzyme but enabled the bovine proteasome to accommodate either the constitutive or the inducible subunits. A novel N-terminal nucleophile hydrolase activity was proposed for the beta 7 subunit. We also determined the site of the nuclear localization signals in the molecule. A model of the immunoproteasome was predicted from this constitutive structure.

The proteasome immunosubunits, PA28 and ER‐aminopeptidase 1 protect melanoma cells from efficient MART‐126‐35‐specific T‐cell recognition

“…We here demonstrate that in addition, the third IFN-γ−inducible IP subunit β2i/MECL-1 contributes to reduced MART-1 epitope generation due to impaired N-and C-terminal cleavage site usage. Reduced N-terminal cleavage may be explained by the reduced β1-peptidylglutamyl peptide-hydrolyzing activity due to β1i/LMP2 (and a polar character of its substrate-binding pocket S1) incorporation into 20S proteasome complexes and for β2i/MECL-1 in that it also contributes to the formation of the S1 pocket of β1i/LMP2 [32,33]. Although concerted PA28α/β and IP-expression is increased following IFN-γ induction, PA28 can also be detected in tissues and cells with low IFN sensitivity such as muscle and erythrocytes that express SP complexes instead of IPs [34,35].…”

“…These data show that ERAP1/2 negatively contributes to MART-1 26-35 epitope generation and that in particular, ERAP1 is responsible for the impairment of epitope presentation by IFN-γ. To study the underlying molecular mechanism, we next performed in vitro trimming experiments using recombinant ERAPs in combination with the N-terminally extended 14-mer epitope precursor peptide MART-1 [22][23][24][25][26][27][28][29][30][31][32][33][34][35] . As shown by reversed-phase HPLC analyses, ERAP1 efficiently trimmed the MART-1 22-35 precursor peptide.…”

“…In vitro studies employing high-affinity MART-1-specific CD8 + T-cell clones demonstrated that some melanoma cell lines, despite expression of MART-1 and the corresponding HLA allele, were only barely recognized by specific T cells [3]. While different variants of the MART-1 epitope (including the MART-1 [26][27][28][29][30][31][32][33][34][35] 10-mer and the MART-1 27-35 9-mer) have been identified, only the 9-mer epitope can be eluted in significant amounts from the cell surface of melanoma cells [4]. This suggests that mechanisms related to antigen processing interfere with efficient generation of the 10-mer MART-1 [26][27][28][29][30][31][32][33][34][35] epitope in tumor cells.…”

“…It has been demonstrated that MART-1 [26][27][28][29][30][31][32][33][34][35] epitope processing is controlled by the proteasome, the major proteolytic machinery of the cytosol [5]. The standard proteasome (SP) contains the catalytic subunits β1, β2, and β5, whereas exposure of cells to IFN-γ enhances expression and incorporation of the immunosubunits β1i/LMP2 (LMP, low molecular weight protein), β2i/MECL-1 (multicatalytic endopeptidase complex-like 1), and β5i/LMP7 into nascent proteasome complexes.…”

Computational Prediction of the Specificities of Proteasome Interaction with Antigen Protein

“…But this paper has the opposite result, and this is probably caused by the difference of structures of different species 20S proteasome. Unno et al has confirmed that the  subunits of human proteasome have some obvious structure differences with the  subunits of yeast proteasome, and the  subunits of human proteasome also have some structure differences with the  subunits of yeast proteasome (23).…”