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Related Concept Videos

The Proteasome02:18

The Proteasome

Eukaryotic cells can degrade proteins through several pathways. One of the most important amongst these is the ubiquitin-proteasome pathway. It helps the cell eliminate the misfolded, damaged, or unwarranted cytoplasmic proteins in a highly specific manner.
In this pathway, the target proteins are first tagged with small proteins called ubiquitin. A series of enzymes carry out the ubiquitination of the target proteins - E1 (ubiquitin-activating enzyme), E2 (ubiquitin-conjugating enzyme), and E3...
The Proteasome01:13

The Proteasome

Eukaryotic cells can degrade proteins through several pathways. One of the most important among these is the ubiquitin-proteasome pathway. It helps the cell eliminate the misfolded, damaged, or unwarranted cytoplasmic proteins in a highly specific manner.
In this pathway, the target proteins are first tagged with small proteins called ubiquitin. This involves participation of a series of enzymes including— E1 (ubiquitin-activating enzyme), E2 (ubiquitin-conjugating enzyme), and E3 (ubiquitin...
The Proteasome Structure01:17

The Proteasome Structure

The ubiquitin-proteasome pathway is a well-known mechanism utilized by eukaryotic cells to remove cytoplasmic proteins that are misfolded, damaged, or no longer needed. In this pathway, the protein that needs to be eliminated undergoes a process called ubiquitination, where a chain of ubiquitin molecules is attached to the 48th lysine residue of the target protein. This ubiquitin modification helps the proteasome distinguish between a target protein and a healthy protein.
The proteasome is an...
Translocation of Proteins into the Mitochondria01:19

Translocation of Proteins into the Mitochondria

Mitochondrial precursors are translocated to the internal subcompartments via independent mechanisms involving distinct protein machineries called translocases.
Sorting of outer membrane proteins:
Mitochondrial outer membrane proteins are of two types: the transmembrane, beta-barrel porins, and the membrane-anchored, alpha-helical proteins. Beta-barrel porin precursors are translocated by the TOM complex and inserted into the outer mitochondrial membrane by the SAM complex. In contrast,...
Mitochondrial Precursor Proteins01:39

Mitochondrial Precursor Proteins

Mitochondrial precursors are partially unfolded or loosely folded polypeptide chains. Newly synthesized precursors are inhibited from spontaneously folding into their native conformation by the cytosolic chaperones, heat shock proteins 70 (Hsp70), and mitochondrial import stimulation factors (MSFs). Precursors bound to MSFs are guided to the TOM70-TOM37 receptors, while precursors bound to Hsp70  chaperones are targetted to TOM20-TOM22 receptor complexes.
Most of the mitochondrial precursors...
Conservative Site-specific Recombination and Phase Variation02:53

Conservative Site-specific Recombination and Phase Variation

Because the DNA segments are cut and reorganized in a direction-specific manner, site-specific recombination has emerged as an efficient genetic engineering technique. Flippase and Cyclization recombinases or Flp and Cre, respectively, are two members of the tyrosine recombinase family derived from bacteriophages, that are used to mediate site-specific DNA insertions, deletions, and targeted expression of proteins in mammalian cell lines.
The recognition sites for Cre recombinase called LoxP...

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A High-Throughput Luciferase Assay to Evaluate Proteolysis of the Single-Turnover Protease PCSK9
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Antitopes define preferential proteasomal cleavage site usage.

Britta Strehl1, Kathrin Textoris-Taube, Sandra Jäkel

  • 1Institut für Biochemie and Klinik für Kardiologie und Pulmologie, Charité-Universitätsmedizin, and Max-Planck-Institut für Infektionsbiologie, Berlin, Germany.

The Journal of Biological Chemistry
|April 22, 2008
PubMed
Summary

Interferon gamma-induced immunoproteasomes enhance antigenic peptide generation by altering proteasomal cleavage specificity, not just substrate turnover. This study introduces a novel "antitope" method to quantify peptide generation for major histocompatibility class I presentation.

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Published on: May 25, 2018

Area of Science:

  • Immunology
  • Molecular Biology
  • Proteomics

Background:

  • Proteasomes degrade proteins, generating peptides for Major Histocompatibility Complex (MHC) class I presentation.
  • Interferon gamma-induced immunoproteasomes enhance antigenic peptide generation, but the mechanism (substrate turnover vs. cleavage specificity) remains unclear.

Purpose of the Study:

  • To elucidate whether enhanced immunoproteasome activity is due to increased substrate processing or altered cleavage specificity.
  • To introduce and validate a novel method for quantifying proteasomal peptide generation for MHC class I presentation.

Main Methods:

  • Development and application of the "antitope" technique as a substrate-specific internal standard for peptide quantification.
  • Analysis of proteasomal cleavage preferences for specific T-cell epitopes, including LLO(296-304) and Coxsackievirus-derived epitopes.

Main Results:

  • Demonstrated that enhanced immunoproteasome-dependent presentation of the LLO(296-304) epitope is caused by altered cleavage preferences.
  • Validated the antitope method's applicability to other MHC class I epitopes, including those from Coxsackievirus.

Conclusions:

  • Immunoproteasome-mediated enhancement of antigenic peptide generation for MHC class I presentation is primarily driven by altered proteasomal cleavage specificity.
  • The antitope method provides a robust tool for quantifying peptide generation and studying proteasome function in antigen presentation.