Cell Signaling Technology

Product Pathways - Protein Stability

PSMB5 Antibody #11903

Applications Reactivity Sensitivity MW (kDa) Source
W H M R Mk Endogenous 22 Rabbit

Applications Key:  W=Western Blotting
Reactivity Key:  H=Human  M=Mouse  R=Rat  Mk=Monkey
Species cross-reactivity is determined by western blot. Species enclosed in parentheses are predicted to react based on 100% sequence homology.

Protocols

Specificity / Sensitivity

PSMB5 Antibody recognizes endogenous levels of total PSMB5 protein. Based upon sequence alignment, this antibody is predicted to react with precursor and mature forms of PSMB5. This antibody does not cross-react with PSMB8.

Source / Purification

Polyclonal antibodies are produced by immunizing animals with a synthetic peptide corresponding to residues near the carboxy terminus of human PSMB5 protein. Antibodies are purified by protein A and peptide affinity chromatography.

Western Blotting

Western Blotting

Western blot analysis of extracts from various cell lines using PSMB5 Antibody.

Western Blotting

Western Blotting

Western blot analysis of extracts from 293T cells, mock transfected (-) or transfected with constructs expressing Myc/DDK-tagged full-length human PSMB5 (hPSMB5-Myc/DDK; +) and Myc/DDK-tagged full-length human PSMB8 (hPSMB8-Myc/DDK; +), using PSMB5 Antibody (upper) or DYKDDDDK Tag Antibody #2368 (lower).

Western Blotting

Western Blotting

Western blot analysis of extracts from HeLa cells, untreated (-) or treated with Human Interferon-γ (hIFN-γ) #8901 (100 ng/ml, 72hr; +), using PSMB5 Antibody (upper) or GAPDH (D16H11) XP® Rabbit mAb #5174 (lower).


Background

The 26S proteasome is a highly abundant ~2 MDa complex that serves as the proteolytic arm of the ubiquitin-proteasome system. It consists largely of two sub-complexes, the 19S regulatory particle (RP) and the 20S catalytic core particle (CP), and in many cases two RPs cap either end of a CP. The CP is made of two stacked β-rings that contain the catalytic sites, each of which is made of seven subunits (β1-7), flanked on either side by two α-rings, which are also made of seven subunits each (α1-7). Thus, the structure of the 20S CP is α1-7β1-7β1-7α1-7. The RP includes a base and a lid. The base, in part, is composed of a hexametric ring of ATPases that function to unfold the substrate and open the gate of the interlacing N-terminal segments of the α-subunits, thus allowing entry of the unfolded substrate into the catalytic chamber. The lid is predominantly involved in specific recognition of the ubiquitin signal (1). In addition to the 19S cap, other proteins and complexes, such as proteasome activator 28 (PA28/11S), bind to the end of the 20S cylinder and activate it by facilitating opening of the gate. Furthermore, proteasome-associated DUBs and E3s can remodel substrate-anchored polyubiquitin chains, which may modulate their susceptibility to degradation (2).

The core particle performs three types of catalytic activities inside its chamber: chymotrypsin-like, trypsin-like, and caspase-like activities, which are provided by the constitutively expressed PSMB5 (β5/MB1/X/LMPX/Macropain epsilon chain), PSMB7 (β2/Z/Macropain chain Z) and PSMB6 (β1/Y/LMPY/Macropain delta chain) subunits, respectively. These catalytic subunits belong to the N-terminal nucleophile (Ntn) hydrolase family and are characterized by an unusual, essentially single-residue active site: The N-terminal threonine of each proteolytic subunit provides both the catalytic nucleophile (on its side chain) and the primary proton acceptor (on the main chain N-terminus). The catalytic β-subunits are synthesized with N-terminal propeptides, which are removed at the final step of proteasome biogenesis by limited proteolysis to expose the catalytic threonine residues (3). In immune responsive cells the constitutively expressed PSMB6, PSMB7, and PSMB5 subunits are replaced by three highly homologous induced β-subunits: PSMB9 (β1i/LMP2/RING12), PSMB10 (β2i/MECL-1/LMP10) and PSMB8 (β5i/LMP7/RING10), respectively, to form the immunoproteasome that has higher chymotrypsin-like and trypsin-like activities known to be favorable for antigen processing (4,5). PSMB5 is downregulated at the protein level by IFN-γ and replaced by PSMB8 in order to remodel the proteolytic specificity of the proteasome for more appropriate immunological processing of endogenous antigens (6-8). PSMB5 is also one of the predominant targets of bortezomib, an inhibitor of the chymotrypsin-like activity of the proteasome (9).

  1. Finley, D. (2009) Annu Rev Biochem 78, 477-513.
  2. Lee, M.J. et al. (2011) Mol Cell Proteomics 10, R110.003871.
  3. Murata, S. et al. (2009) Nat Rev Mol Cell Biol 10, 104-15.
  4. Boes, B. et al. (1994) J Exp Med 179, 901-9.
  5. Cardozo, C. and Kohanski, R.A. (1998) J Biol Chem 273, 16764-70.
  6. Akiyama, K. et al. (1994) Science 265, 1231-4.
  7. Akiyama, K. et al. (1994) FEBS Lett 343, 85-8.
  8. Gaczynska, M. et al. (1996) J Biol Chem 271, 17275-80.
  9. Oerlemans, R. et al. (2008) Blood 112, 2489-99.

Application References

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Companion Products

For Research Use Only. Not For Use In Diagnostic Procedures.

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