The human multicatalytic proteinase: affinity purification using a monoclonal antibody

Hendil, Klavs B.; Uerkvitz, Wolfgang

doi:10.1016/0165-022x(91)90028-u

Cited by 44 publications

(26 citation statements)

References 17 publications

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“…This result indicates that proteasome β1i-β5i is less active at destroying the antigenic peptide. As a result, fragments with the proper N terminus or C terminus of the antigenic The proportions of four proteasomes types were determined by two different sandwich ELISAs, indicated by* and † , both using MCP21 as capture antibody (39). *This ELISA quantified all proteasomes remaining after depletions as described in Fig.…”

Section: Resultsmentioning

confidence: 99%

Two abundant proteasome subtypes that uniquely process some antigens presented by HLA class I molecules

Guillaume

Chapiro

Stroobant

et al. 2010

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

190

273

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Most antigenic peptides presented by MHC class I molecules result from the degradation of intracellular proteins by the proteasome. In lymphoid tissues and cells exposed to IFNγ, the standard proteasome is replaced by the immunoproteasome, in which all of the standard catalytic subunits β1, β2, and β5 are replaced by their inducible counterparts β1i, β2i, and β5i, which have different cleavage specificities. The immunoproteasome thereby shapes the repertoire of antigenic peptides. The existence of additional forms of proteasomes bearing a mixed assortment of standard and inducible catalytic subunits has been suggested. Using a new set of unique subunit-specific antibodies, we have now isolated, quantified, and characterized human proteasomes that are intermediate between the standard proteasome and the immunoproteasome. They contain only one (β5i) or two (β1i and β5i) of the three inducible catalytic subunits of the immunoproteasome. These intermediate proteasomes represent between one-third and one-half of the proteasome content of human liver, colon, small intestine, and kidney. They are also present in human tumor cells and dendritic cells. We identified two tumor antigens of clinical interest that are processed exclusively either by intermediate proteasomes β5i ) or by intermediate proteasomes β1i-β5i (MAGE-A10 [254][255][256][257][258][259][260][261][262] ). The existence of these intermediate proteasomes broadens the repertoire of antigens presented to CD8 T cells and implies that the antigens presented by a given cell depend on their proteasome content.antigen processing | antigenic peptide | immunoproteasome | tumor antigen | MAGE

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Section: Resultsmentioning

confidence: 99%

Two abundant proteasome subtypes that uniquely process some antigens presented by HLA class I molecules

Guillaume

Chapiro

Stroobant

et al. 2010

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

190

273

View full text Add to dashboard Cite

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“…Thioflavin S and Congo Red were from Sigma. Human 20 S proteasome was isolated from human erythrocytes by affinity chromatography on immobilized MCP21 monoclonal antibody (26). The specific activities of the 20 S proteasome are as follows: chymotrypsin-like activity, 4.0 nmol/min/mg protein using succinylAla-Ala-Phe-7-amido-4-methylcoumarin; trypsin-like activity, 5.1 nmol/ min/mg protein using Z-Ala-Ala-Arg-7-amido-4-methylcoumarine; caspase-like activity, 1.9 nmol/min/mg protein using Z-Leu-Leu-Glu-␤-naphthylamide.…”

Section: Methodsmentioning

confidence: 99%

“…First, the purified 20 S proteasome (24) was from a commercial source and purified by ammonium sulfate precipitation and several chromatographic steps. In contrast, the 20 S proteasome preparation used in the present study was isolated by a milder procedure involving immunoaffinity chromatography (26). We have experienced with one batch of purified 20 S proteasome stored for prolonged time at Ϫ20°C that it lost the majority of its hydrolytic activity.…”

Section: ␣-Synuclein Inhibits the 20 S Proteasomementioning

confidence: 99%

Proteasomal Inhibition by α-Synuclein Filaments and Oligomers

Lindersson

Beedholm

Højrup

et al. 2004

Journal of Biological Chemistry

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366

310

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A unifying feature of many neurodegenerative disorders is the accumulation of polyubiquitinated protein inclusions in dystrophic neurons, e.g. containing ␣-synuclein, which is suggestive of an insufficient proteasomal activity. We demonstrate that ␣-synuclein and 20 S proteasome components co-localize in Lewy bodies and show that subunits from 20 S proteasome particles, in contrast to subunits of the 19 S regulatory complex, bind efficiently to aggregated filamentous but not monomeric ␣-synuclein. Proteasome binding to insoluble ␣-synuclein filaments and soluble ␣-synuclein oligomers results in marked inhibition of its chymotrypsin-like hydrolytic activity through a non-competitive mechanism that is mimicked by model amyloid-A␤ peptide aggregates. Endogenous ligands of aggregated ␣-synuclein like heat shock protein 70 and glyceraldehyde-6-phosphate dehydrogenase bind filaments and inhibit their anti-proteasomal activity. The inhibitory effect of amyloid aggregates may thus be amenable to modulation by endogenous chaperones and possibly accessible for therapeutic intervention.

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“…Cleared supernatants were desalted on a Hiprep 26/10 desalting column (GE Healthcare) equilibrated with buffer. Proteasomes were first purified by immunoaffinity chromatography on CNBr-activated 4B Sepharose beads (63 ml; Amersham Pharmacia Biotech) coupled to mAb MCP21, which is directed against the a2 subunit of the proteasome (16). Proteasomes were eluted with 20 mM Tris-HCl and 3 M NaCl.…”

Section: Proteasome Purificationmentioning

confidence: 99%

Analysis of the Processing of Seven Human Tumor Antigens by Intermediate Proteasomes

Guillaume

Stroobant

Bousquet

et al. 2012

The Journal of Immunology

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We recently described two proteasome subtypes that are intermediate between the standard proteasome and the immunoproteasome. They contain only one (β5i) or two (β1i and β5i) of the three inducible catalytic subunits of the immunoproteasome. They are present in tumor cells and abundant in normal human tissues. We described two tumor antigenic peptides that are uniquely produced by these intermediate proteasomes. In this work, we studied the production by intermediate proteasomes of tumor antigenic peptides known to be produced exclusively by the immunoproteasome (MAGE-A3114–122, MAGE-C242–50, MAGE-C2336–344) or the standard proteasome (Melan-A26–35, tyrosinase369–377, gp100209–217). We observed that intermediate proteasomes efficiently produced the former peptides, but not the latter. Two peptides from the first group were equally produced by both intermediate proteasomes, whereas MAGE-C2336–344 was only produced by intermediate proteasome β1i-β5i. Those results explain the recognition of tumor cells devoid of immunoproteasome by CTL recognizing peptides not produced by the standard proteasome. We also describe a third antigenic peptide that is produced exclusively by an intermediate proteasome: peptide MAGE-C2191–200 is produced only by intermediate proteasome β1i-β5i. Analyzing in vitro digests, we observed that the lack of production by a given proteasome usually results from destruction of the antigenic peptide by internal cleavage. Interestingly, we observed that the immunoproteasome and the intermediate proteasomes fail to cleave between hydrophobic residues, despite a higher chymotrypsin-like activity measured on fluorogenic substrates. Altogether, our results indicate that the repertoire of peptides produced by intermediate proteasomes largely matches the repertoire produced by the immunoproteasome, but also contains additional peptides.

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The human multicatalytic proteinase: affinity purification using a monoclonal antibody

Cited by 44 publications

References 17 publications

Two abundant proteasome subtypes that uniquely process some antigens presented by HLA class I molecules

Two abundant proteasome subtypes that uniquely process some antigens presented by HLA class I molecules

Proteasomal Inhibition by α-Synuclein Filaments and Oligomers

Analysis of the Processing of Seven Human Tumor Antigens by Intermediate Proteasomes

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