Proteases proteasome

If RTA unfolds to exploit the target cell ERAD pathway, then it must refold or otherwise escape the normal fate of misfolded cytosolic proteins, which typically includes degradation by ATP-dependent proteases (proteasome). One protective mechanism for avoiding degradation appears to be an inherently low number of exposed lysine residues on RTA (2 per monomer) exposed lysine residues are a primary target for ubiquitination, and the presence of ubiquitin routes misfolded proteins to the proteasome (Decks et al., 2002). 

Baumeister W, Walz J, Zuhl F and Seemuller E 1998 The proteasome paradigm of a self-oompartmentalizing protease Cell 92 367-80... 

Several aryl esters of 6-chloromethyl-2-oxo-2//-l -benzopyran-3-carboxylic acid act as human Lon protease inhibitors (alternate substrate inhibitors)46 without having any effect on the 20S proteasome. Proteasomes are the major agents of protein turnover and the breakdown of oxidized proteins in the cytosol and nucleus of eukaryotic cells,47 whereas Lon protease seems to play a major role in the elimination of oxidatively modified proteins in the mitochondrial matrix. The coumarin derivatives are potentially useful tools for investigating the various biological roles of Lon protease without interfering with the proteasome inhibition. 

Bayot, A. Basse, N. Lee, I. Gareil, M. Pirotte, B. Bulteau, A. L. Friguet, B. Reboud-Ravaux, M. Towards the control of intracellular protein turnover mitochondrial Lon protease inhibitors versus proteasome inhibitors. Biochimie 2008, 90, 260-269. 

E. Seemiiller, A. Lupas, D. Stock, J. Lowe, R. Huber, W. Baumeister, Proteasome from Thermoplasma Acidophilum A Threonine Protease , Science 1995, 268, 579-582. 

Three other components that my laboratory has identified and partially purified from Fraction 2 of reticulocytes, termed CF1-CF3, are involved in the degradation of proteins ligated to ubiquitin [24]. These are apparently subcomplexes of the 26S proteasome, a large ATP-dependent protease complex first described by Re-chsteiner and co-workers [25], CF3 is identical to the 20S proteasome core particle [26], while CFl and CF2 may be similar to the base and lid subcomplexes of the 19S regulatory particle of the 26S proteasome, described more recently by the Finley laboratory [27], In hindsight, the reason for finding subcomplexes, rather than the complete 26S complex in Fraction 2 was technical we have routinely prepared Fraction 2 from ATP-depleted reticulocytes [20], under which conditions the 26S proteasome dissociates to its subcomplexes. We found that incubation of the three subcomplexes in the presence of ATP promotes their assembly to the 26S proteasome [24, 26]. The role of ATP in the assembly of the 26S proteasome complex remains unknown. 

Yao, T. and Cohen, R. E. A cryptic protease couples deubiquitination and degradation by the proteasome. 

Whereas standard proteases use serine, cysteine, aspartate, or metals to cleave peptide bonds, the proteasome employs an unusual catalytic mechanism. N-terminal threonine residues are generated by self-removal of short peptide extensions from the active yS-subunits and act as nucleophiles during peptide-bond hydrolysis [23]. Given its unusual catalytic mechanism, it is not surprising that there are highly specific inhibitors of the proteasome. The fungal metabolite lactacystin and the bacterial product epoxomicin covalently modify the active-site threonines and in-... 

Both the 26S proteasome and the RC hydrolyze all four nucleotide triphosphates, with ATP and CTP preferred over GTP and UTP [58]. Although ATP hydrolysis is required for conjugate degradation, the two processes are not strictly coupled. Complete inhibition of the peptidase activity of the 26S proteasome by calpain inhibitor I has little effect on the ATPase activity of the enzyme. The nucleotidase activities of the RC and the 26S proteasome closely resemble those of E. coli Lon protease, which is composed of identical subunits that possess both proteolytic and nucleotidase activities in the same polypeptide chain. Like the regulatory complex and 26S proteasome, Lon hydrolyzes all four ribonucleotide triphosphates, but not ADP or AMP [18]. 

The 20S proteasome is a latent protease owing to the barrier imposed by the a-subunit rings on peptide entry. Consequently, a readily measured activity of the RC is activation of fluorogenic peptide hydrolysis by the 20S proteasome. The extent of activation is generally found to be in the range 3- to 20-fold [63]. Activation is relatively uniform for all three proteasome catalytic subunits and presumably reflects opening by the attached RC of a channel leading to the proteasome s central chamber. 

Kessel, M. et al. Homology in structural organization between E. coli ClpAP protease the eukaryotic 26 S proteasome. J Mol Biol 1995, 250, 587-594. 

ATP-dependent proteases are complex proteolytic machines. They are present in eubacteria, archaebacteria, in eukaryotic organelles and, as the 20S or 26S proteasome, in the eukaryotic cytosol and nucleoplasm. The activators of all known ATP-dependent proteases are related. They all contain an AAA(+) ATPase domain as a module (Neuwald et al. 1999) and are thought to assemble into hexameric particles or, in the case of 26S proteasomes, are present in six variants in the 19S activators (Glickman et al. 1999). Like the ATPases, the proteolytic components of the ATP-dependent proteases form higher order complexes, but unlike for the ATPases, the symmetry of the protease assemblies varies, and the folds of the subunits need not be related. ClpP is a serine protease, FtsH a metalloprotease, and HslV and the proteasomes from archaebacteria and eubacteria are threomne proteases. 

The bacterial protease HslVU is unique in two respects at present, it is the only ATP-dependent protease to have atomic coordinates of the full complex determined secondly, and in contrast to all other bacterial ATP-dependent proteases, it contains a proteolytic core that is related to the 20S proteolytic core of archaebacterial and eukaryotic proteasomes. The following sections summarize our understanding of HslVU biochemistry, crystallography, and enzymology and end with some speculation on the implications of these results for other ATP-dependent proteases. 

Rohewild, M., Coux, O., Huang, H. C., Moeeschell, R. P., Soon, J. Y., Seol, J. H., Chung, C. H., and Goldberg, A. L. HslV-HslU A novel ATP-dependent protease complex in Fscherichia coli related to the eukaryotic proteasome. Proc. Natl. Acad. Sci. USA 1996, 93, 5808-5813. 

Rohewild, M., Peeieee, G., Santaeius, U., Muller, S. A., Huang, H. C., Fngel, A., Baumeistee, W., and Goldberg, A. L. The ATP-dependent HslVU protease from Escherichia coli is a four-ring structure resembling the proteasome. Nat Struct. Biol 1997, 4, 133-139. 

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