Ubiquitin-activating enzymes

Ubiquitin tags proteins for protein degradation. The ubiquitination requires three different enzymatic activities, a ubiquitin-activating enzyme (El), a ubiquitin-conjugating enzyme (E2 or Ubc) and a ubiquitin ligase (E3). The action of all three enzymes leads to the establishment of a poly-ubiquitin chain on target proteins which are then recognized and proteolyzed by the 26S proteasome. 

The natural product panepophenanthrin (6/1-170), isolated in 2002 from the fermented broth of the mushroom strain Panus radus IFO 8994 [90], is the first example of an inhibitor of the ubiquitin-activating enzyme [91]. Retrosynthetic analysis based on a biomimetic analysis led to the conjugated diene 6/1-172 by a retro-Diels-Alder reaction via the hemiacetal 6/1-171. Further disconnections of 6/1-172 produces the vinyl stannane 6/1-173 and the vinyl bromide 6/1-174 [92]. 

Ubiquitin-El Ubiquitin-activating enzyme UbL Ubiquitin-like protein... 

A) Schematic comparison between MoeB, the ubiquitin-activating enzyme (UBAl) and the heterodimeric (APPBP1-UBA3) NEDD8 activator based on PSI-BLAST [54] sequence analyses. The numbers represent the sequence identities between the first 170-180 residues of MoeB (shaded) and the corresponding regions in... 

Haas, A. L. and Rose, 1. A. The mechanism of ubiquitin activating enzyme. A kinetic and equilibrium analysis, J Biol Chem 1982, 257, 10329-10337. 

Fig. 5.1. The ubiquitin-conjugation pathway. Steps in ubiquitin activation and substrate modification. El, ubiquitin activating enzyme E2, ubiquitin-conjugating enzyme E3, ubiquitin-protein ligase. Atoms involved in the thiol ester and amide bonds are shown.

Heeshko, A., and Rose, 1. A. Ubiquitin-activating enzyme. Mechanism and role in protein-ubiquitin conjugation. J. Biol. Chem. 1982, 257, 2543-48. 

Fig. 12.1. Domain scheme of selected proteins with internal ubiquitin-like domains. Ubiquitin-like domains are indicated by black boxes. Other domains are abbreviated as follows ThiF, NAD-binding domain in ubiq-uitin activating enzymes UAct, 2nd conserved domain in ubiquitin activating enzymes ...

Increasing evidence indicates that accumulation of aberrant or misfolded proteins, protofibril formation, ubiquitin-proteasome system dysfunction, and the direct or indirect consequences of abnormal protein aggregation and accumulation represent deleterious events linked to neurodegeneration (255,256). Ubiquitination is an essential cellular process affected by a multienzyme cascade involving Els (ubiquitin-activating enzymes), E2s (ubiquitin-conjugation enzymes or UBCs), and E3s (ubiquitin-protein Ugases) (12,257) (see Fig. 10.4). 

Ciechanover, A., Elias, S., Heller, H. and Hershko, A. (1982). Covalent affinity purification of ubiquitin activating enzyme. J. Biol. Chem. 257, 2537-2542. 

Finley, D., Ciechanover, A., and Varshavsky, A. (1984). Thermolability of ubiquitin-activating enzyme from the mammalian cell cycle mutant ts85. Cell 37, 43-55. 

Ubiquitin-Activating Enzyme, Ubiquitin-Conjugating Enzymes... 

Fig. 1. A schematic diagram outlining the hierarchic structure of the ubiquitin system. In an ATP-dependent manner a thioester bond is formed between the C-terminus of ubiquitin and an internal cystein residue of the ubiquitin-activating enzyme. Subsequently, ubiquitin is transferred to a member of the family of ubiquitin-conjugating enzymes, which are also able to form a thioester bond with ubiquitin. The third class of enzymes, the ubiquitin ligases, direct ubiquitin to the proteolytic substrates. Different families of this class of enzymes are known, some of which are also able to form a thioester intermediate with ubiquitin (HECT-domain ligases). The final ubiquitin-substrate linkage is an isopeptide bond between the C-terminus of ubiquitin and internal lysine residues in the substrate proteins...

In mammalian cells the El ubiquitin-activating enzyme exists in two isoforms, Ela (110 kDa) and Elb (117 kDa), which are derived from a single gene and mRNA (Cook and Chock 1992, Handley Gearhart et al. 1994). The isoform Ela is predominantly found in the nucleus and has been shown to harbor a functional nuclear localization sequence (NLS) required for nuclear targeting and phosphorylation. In contrast, Elb lacks the NLS, is not phos-phorylated and localized in the cytoplasm (Handley-Gearhart et al. 1994 Stephen et al. 1997). Phosphorylation of Ela was demonstrated to occur in a cell cycle-dependent manner, being maximal in G2 phase (Stephen et al. 

Cook )C, Chock PB (1992) Isoforms of mammalian ubiquitin-activating enzyme. Biol Chem 267 24315-24321... 

Stephen AG, Trausch Azar JS, Ciechanover A, Schwartz AL (1996) The ubiquitin-activating enzyme El is phosphorylated and localized to the nucleus in a cell cycle-dependent manner. Biol Chem 271 15608-15614 Stephen AG, Trausch Azar JS, Handley Gearhart PM, Ciechanover A, Schwartz AL (1997) Identification of a region within the ubiquitin-activating enzyme required for nuclear targeting and phosphorjdation.) Biol Chem 272 10895-10903 Sternsdorf T, Jensen K, Will H (1997) Evidence for covalent modification of the nuclear dot-associated proteins PML and SplOO by PICl/SUMO-1. J Cell Biol 139 1621-1634... 

In an initial reaction ubiquitin is activated by forming a reactive thiolester with an SH-group of the ubiquitin-activating enzyme El. This step requires ATP and consists of an intermediate formation of ubiquitin adenylate followed by the binding of ubiquitin to a Cys residue of El in a thiolester linkage, with the release of PPi and AMP. 

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