Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:P62988 (Ubiquitin)
 4,326 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Ubiquitin plays essential roles in various cellular processes; therefore, it is of keen interest to study the structure-function relationship of ubiquitin itself. We investigated the modification of Lys(6) of ubiquitin and its physiological consequences. Mass spectrometry-based peptide mapping and N-terminal sequencing demonstrated that, of the 7 Lys residues in ubiquitin, Lys(6) was the most readily labeled with sulfosuccinimidobiotin. Lys(6)-biotinylated ubiquitin was incorporated into high molecular mass ubiquitin conjugates as efficiently as unmodified ubiquitin. However, Lys(6)-biotinylated ubiquitin inhibited ubiquitin-dependent proteolysis, as conjugates formed with Lys(6)-biotinylated ubiquitin were resistant to proteasomal degradation. Ubiquitins with a mutation of Lys(6) had similar phenotypes as Lys(6)-biotinylated ubiquitin. Lys(6) mutant ubiquitins (K6A, K6R, and K6W) also inhibited ATP-dependent proteolysis and caused accumulation of ubiquitin conjugates. Conjugates formed with K6W mutant ubiquitin were also resistant to proteasomal degradation. The dominant-negative effect of Lys(6)-modified ubiquitin was further demonstrated in intact cells. Overexpression of K6W mutant ubiquitin resulted in accumulation of intracellular ubiquitin conjugates, stabilization of typical substrates for ubiquitin-dependent proteolysis, and enhanced susceptibility to oxidative stress. Taken together, these results show that Lys(6)-modified ubiquitin is a potent and specific inhibitor of ubiquitin-mediated protein degradation.
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PMID:Lys6-modified ubiquitin inhibits ubiquitin-dependent protein degradation. 1579 May 62

Eukaryotic elongation factor-2 kinase (eEF-2 kinase) is a highly conserved calcium/calmodulin-dependent enzyme involved in the regulation of protein translation and cell proliferation. Rapid changes in the activity and abundance of eEF-2 kinase have been observed on growth stimulation, and increased enzyme activity is characteristic of malignant cell growth. Yet the mechanism for controlling the turnover of this kinase is unknown. The ubiquitin-proteasome pathway regulates the degradation of many cellular proteins, including transcription factors, cell cycle regulators, and signal transduction proteins. Therefore, we determined whether the ubiquitin-proteasome pathway regulates the turnover of eEF-2 kinase. We found that eEF-2 kinase was a relatively short-lived protein with a half-life of less than 6 hours. eEF-2 kinase was ubiquitinated in vivo as determined by coimmunoprecipitation and polyubiquitin affinity matrix. Incubation of purified eEF-2 kinase with a source of ubiquitination enzymes (rabbit reticulocyte lysate), purified ubiquitin, and ATP revealed the presence of increasing molecular weight species of ubiquitinated eEF-2 kinase. Treatment of cells with MG132, a proteasome inhibitor, inhibited eEF-2 kinase degradation and induced the accumulation of polyubiquitinated forms of the enzyme, resulting in an increase in its half-life. These results suggest involvement of the proteasome in the turnover of the ubiquitinated kinase. Because eEF-2 kinase is chaperoned by heat shock protein 90 (Hsp90), we next determined if disruption of the Hsp90-eEF-2 kinase complex promoted degradation of the kinase. Treatment of cells with geldanamycin, an Hsp90 inhibitor, enhanced ubiquitination of eEF-2 kinase and decreased the half-life of the kinase to less than 2 hours. These results indicate that cellular levels of eEF-2 kinase are maintained by a balance between association with Hsp90 and degradation by the ubiquitin-proteasome pathway. In conclusion, these data show that the turnover of eEF-2 kinase is regulated by the ubiquitin-proteasome pathway and, therefore, modulating the ubiquitination of eEF-2 kinase might control the abundance of this enzyme and have implications in the treatment of certain forms of cancer.
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PMID:Identification of the ubiquitin-proteasome pathway in the regulation of the stability of eukaryotic elongation factor-2 kinase. 1586 77

ATP hydrolysis is required for degradation of polyubiquitinated proteins by the 26S proteasome but is thought to play no role in proteasomal stability during the catalytic cycle. In contrast to this view, we report that ATP hydrolysis triggers rapid dissociation of the 19S regulatory particles from immunopurified 26S complexes in a manner coincident with release of the bulk of proteasome-interacting proteins. Strikingly, this mechanism leads to quantitative disassembly of the 19S into subcomplexes and free Rpn10, the polyubiquitin binding subunit. Biochemical reconstitution with purified Sic1, a prototype substrate of the Cdc34/SCF ubiquitin ligase, suggests that substrate degradation is essential for triggering the ATP hydrolysis-dependent dissociation and disassembly of the 19S and that this mechanism leads to release of degradation products. This is the first demonstration that a controlled dissociation of the 19S regulatory particles from the 26S proteasome is part of the mechanism of protein degradation.
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PMID:ATP hydrolysis-dependent disassembly of the 26S proteasome is part of the catalytic cycle. 2967 18

Ubiquitin is highly conserved 76 amino acid protein found in all eukaryotic organisms and ubiquitin-proteasome pathway (UPP) plays a very important role in regulated non-lysosomal ATP dependent protein degradation. This pathway participates in or regulates numerous cellular processes, such as selective protein degradation, cell cycle progression, apoptosis, signal transduction, transcriptional regulation, receptor control by endocytosis, immune response and the processing of antigens. Nevertheless, roles of UPP in virus infection are only beginning to be clarified. Ubiquitin homology has also been found in insect viruses. All viral ubiquitin genes encode an N-terminal ubiquitin sequence and 3-256 amino acids C-terminal peptides. Most of the residues known to be essential for ubiquitin function have been conserved in the viral variant. In Autographa californica nucleopolyhedrovirus (AcMNPV), viral ubiquitin is attached to the inner surface of budded viron membrane by a covalently linked phospholipid and is not essential for viral replication. Currently, insect viruses are the only viruses known to encode ubiquitin. However, ubiquitin also plays a role in the life cycle of other viruses. Host ubiquitin molecules have been found in some plant viruses and other animal viruses. Additionally, Africa swine fever virus (ASFV) encodes a ubiquitin-conjugating enzyme (E2) and a putative causal link between human immunodeficiency virus type 1 (HIV-1) and ubiquitin was established by showing that depletion of the intracellular pool of free ubiquitin inhibits the virus budding. Further analyses indicated that many retroviruses proteins which are required for efficient pinching off the virus bud contain a late domain. The core element of the late domain is a proline-rich motif (PPXY) which mediates the late domain to be ubiquitinated by cellular proteins. Recently, it has been shown that many retroviruses have developed mechanisms to escape the cellular immune response, to facilitate virus replication and to promote virus assembly and budding via host UPP.
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PMID:[Ubiquitin-proteasome pathway and virus infection]. 1596

The 97-kDa molecular chaperone valosin-containing protein (VCP) belongs to a highly conserved AAA family and forms a hexameric structure that is essential for its biological functions. The AAA domain contains highly conserved motifs, the Walker A, Walker B, and the second region of homology (SRH). Although Walker A and B motifs mediate ATP binding and hydrolysis, respectively, the function of the SRH in VCP is not clear. We examined the significance of the SRH in VCP, especially the conserved Arg(359) and Arg(362) in the first AAA domain, D1 and Arg(635) and Arg(638) in the second AAA domain, D2. We show that Arg(359) and Arg(362) in D1 are critical for maintaining the hexameric structure and the ability to bind the polyubiquitin chains. Although the rest of the tested SRH mutants retain the hexameric structure, all of them exhibit severely reduced ATPase activity. Tryptophan fluorescence analysis showed that all of the tested mutants can bind to ATP or ADP. Thus, the reduced ATPase activity likely results from the hampered communications among protomers during hydrolysis. Moreover, when the ATPase-defective mutant R635A or R638A is mixed with the Walker A mutant of D2, the ATPase activity is partially restored, suggesting that Arg(635) and Arg(638) can stimulate the ATPase activity of the neighboring protomer. Interestingly, mutation of Arg(359) and Arg(362) uncouples the inhibitory effect of p47, a VCP co-factor, on the ATPase activity of VCP. Therefore, the Arg residues allow D1 to take on a specific conformation that is required for substrate binding and co-factor communications. Taken together, these results demonstrate that the conserved Arg residues in the SRH of both D1 and D2 play critical roles in communicating the conformational changes required for ATP hydrolysis, and SRH in D1 also contributes to substrate binding and co-factor communications.
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PMID:Multifunctional roles of the conserved Arg residues in the second region of homology of p97/valosin-containing protein. 1621 72

It is increasingly appreciated that failures in the ubiquitin-proteasome system play a pivotal role in the neuropathogenesis of many neurological disorders. This system, involved in protein quality control, should degrade misfolded proteins, but apparently during neuropathogenesis, it is unable to cope with a number of proteins that, by themselves, can consequently accumulate. Ubiquitin is essential for ATP-dependent protein degradation by the proteasome. Ubiquitin+1 (UBB+1) is generated by a dinucleotide deletion (DeltaGU) in UBB mRNA. The aberrant protein has a 19 amino acid extension and has lost the ability to ubiquitinate. Instead of targeting proteins for degradation, it has acquired a dual substrate-inhibitor function; ubiquitinated UBB+1 is a substrate for proteasomal degradation, but can at higher concentrations inhibit, proteasomal degradation. Furthermore, UBB+1 protein accumulates in neurons and glial cells in a disease-specific way, and this event is an indication for proteasomal dysfunction. Many neurological and non-neurological conformational diseases have the accumulation of misfolded proteins and of UBB+1 in common, and this combined accumulation results in the promotion of insoluble protein deposits and neuronal cell death as shown in a cellular model of Huntington's disease.
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PMID:Conformational diseases: an umbrella for various neurological disorders with an impaired ubiquitin-proteasome system. 1622 48

Ubiquitin is a highly conserved, 76-amino acid polypeptide with several important regulatory functions in both plants and animals that all arise from its covalent ligation to other cellular proteins. Here, we demonstrate that higher plants have the capacity to conjugate ubiquitin to other plant proteins in vitro. Using (125)I-labeled human ubiquitin as a substrate, conjugating activities were observed in crude etiolated tissue extracts from all species tested, including oats, rye, barley, corn, zucchini squash, pea, soybean, and sunflower. The reaction has a soluble distribution, is specific for ATP, and requires the protease inhibitor, leupeptin, to protect ubiquitin from inactivation during the assay. Conjugation is inhibited by N-ethylmaleimide and high concentrations of 2-mercaptoethanol suggesting that the mechanism of ubiquitin ligation in plants involves a similar thiolester intermediate to that found in the mammalian pathway. The conjugating activity in etiolated oat extracts is extremely labile with a half-life of about 20 minutes at 30 degrees C. Detectable but low ATP-stimulated, conjugating activities were also observed in extracts from dry seeds and green leaves of oats. In addition to this conjugating activity, crude plant extracts have the capacity to degrade ubiquitin-protein conjugates formed in vitro. These results demonstrate that higher plants contain several of the enzymic activities necessary for ubiquitin's functions and provide a method for assaying ubiquitin conjugation in vitro.
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PMID:Demonstration of ATP-Dependent, Ubiquitin-Conjugating Activities in Higher Plants. 1666 39

Ubiquitin, a key component in an ATP-dependent proteolytic pathway, participates in the response of various eucaryotic organisms to high temperature stress. Our objective was to determine if ubiquitin serves a similar capacity for metabolizing altered proteins in higher plants during stress. Degradation of total proteins was measured, and ubiquitin pools (free versus conjugated) were extracted with an improved protocol from wheat (Triticum aestivum L. cv Len) roots treated at 22, 27, 32, 37, and 42 degrees C for 1 hour and assayed by western blots and radioimmunoassays. Heat-shock protein synthesis was detected by in vivo labeling and autoradiography. Mean half-life of total root proteins decreased from 51 hours at 22 degrees C to 23 hours at 40 degrees C. Ubiquitin pools were extracted better and proteolysis was slowed more by the improved protocol than by a conventional procedure for plant proteins. Amounts of high molecular mass conjugates were elevated and levels of low molecular mass conjugates and free ubiquitin were depressed when roots were treated at 37 or 42 degrees C than at lower temperatures; the same high temperatures also induced synthesis of heat-shock proteins. We concluded that high temperatures increase breakdown of root proteins, which are degraded via the ubiquitin proteolytic pathway. A conjugate with an apparent molecular mass of 23 kilodaltons was tentatively identified as an ubiquitinated histone.
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PMID:Ubiquitin Pool Modulation and Protein Degradation in Wheat Roots during High Temperature Stress. 1666 43

UBE1 is known as the human ubiquitin-activating enzyme (E1), which activates ubiquitin in an ATP-dependent manner. Here, we identified a novel human ubiquitin-activating enzyme referred to as UBE1L2, which also shows specificity for ubiquitin. The UBE1L2 sequence displays a 40% identity to UBE1 and also contains an ATP-binding domain and an active site cysteine conserved among E1 family proteins. UBE1L2 forms a covalent link with ubiquitin in vitro and in vivo, which is sensitive to reducing conditions. In an in vitro polyubiquitylation assay, recombinant UBE1L2 could activate ubiquitin and transfer it onto the ubiquitin-conjugating enzyme UbcH5b. Ubiquitin activated by UBE1L2 could be used for ubiquitylation of p53 by MDM2 and supported the autoubiquitylation of the E3 ubiquitin ligases HectH9 and E6-AP. The UBE1L2 mRNA is most abundantly expressed in the testis, suggesting an organ-specific regulation of ubiquitin activation.
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PMID:UBE1L2, a novel E1 enzyme specific for ubiquitin. 1758 Mar 10

The ubiquitin-proteasome system has come to be known as a vital constituent of mammalian cells. The proteasome is a large nonlysosomal enzyme that acts in concert with an 8.5 kDa polypeptide called ubiquitin and a series of conjugating enzymes, known as E1, E2 and E3, that covalently bind multiple ubiquitin moieties in a polyubiquitin chain to protein substrates in a process called ubiquitylation. The latter process targets protein substrates for unfolding and degradation by the 26S proteasome. This enzyme system specifically recognizes and degrades polyubiquitylated proteins, many of which are key proteins involved in cell cycle regulation, apoptosis, signal transduction, and antigen presentation. The 26S proteasome contains a cylinder-shaped 20S catalytic core that, itself, degrades proteins in an ATP- and ubiquitin-independent manner. The 20S form is actually the predominant enzyme form in mammalian cells. Proteolysis by the constitutive 20S proteasome is vital in removing oxidized, misfolded and otherwise modified proteins. Such degradation is critical as a means of cellular detoxification, as intracellular accumulation of damaged and misfolded proteins is potentially lethal. Studies have shown that inhibition of proteasome activity can lead to cell death. Ethanol and its metabolism cause partial inhibition of the proteasome. This leads to a number of pleiotropic effects that can affect a variety of cellular processes. This critical review describes important aspects of ethanol metabolism and its influence on the proteasome. The review will summarize recent findings on: (1) the interactions between the proteasome and the ethanol metabolizing enzyme, CYP2E1; (2) the dynamics of proteasome inhibition by ethanol in animal models and cultured cells; (3) ethanol-elicited suppression of proteasome activity and its effect on signal transduction; (4) The role of proteasome inhibition in cytokine production by liver cells; and (5) ethanol elicited suppression of peptide hydrolysis and the potential effects on antigen presentation. While the principal focus is on alcohol-induced liver injury, the authors foresee that the findings presented in this review will prompt further research on the role of this proteolytic system in other tissues injured by excessive alcohol consumption.
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PMID:Role of the proteasome in ethanol-induced liver pathology. 1776 Jul 83


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