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Query: UNIPROT:P62988 (Ubiquitin)
 4,326 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Ubiquitin, a 76-residue protein, occurs in cells either free or covalently joined to a variety of protein species, from chromosomal histones to cytoplasmic proteins. Conjugation of ubiquitin to proteolytic substrates is essential for the selective degradation of intracellular proteins in higher eukaryotes. We show here that a protein homologous to human ubiquitin exists in the yeast Saccharomyces cerevisiae, and that yeast extracts conjugate human ubiquitin to a variety of endogenous proteins in an ATP-dependent reaction. We have isolated the S. cerevisiae ubiquitin gene and found it to contain six consecutive ubiquitin-coding repeats in a found it to contain six consecutive ubiquitin-coding repeats in a head-to-tail arrangement. This apparently unique gene organization suggests that yeast ubiquitin is generated by processing of a precursor protein in which several exact repeats of the ubiquitin amino acid sequence are joined directly via Gly-Met peptide bonds between the last and first residues of mature ubiquitin, respectively. Ubiquitin-coding yeast DNA repeats are restricted to a single genomic locus; although the sequenced repeats differ in up to 27 of 228 bases per repeat, they encode identical amino acid sequences. As this predicted amino acid sequence differs in only 3 of 76 residues from that of ubiquitin in higher eukaryotes, ubiquitin is apparently the most conserved of known proteins.
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PMID:The yeast ubiquitin gene: head-to-tail repeats encoding a polyubiquitin precursor protein. 609 20

ts85 cell is a temperature-sensitive mutant of cell cycle, and chromosomal protein uH2A of this mutant disappears at the non-permissive temperature. uH2A in nucleosomes is thought to be synthesized or degradated as follows. H2A + Ubiquitin in equilibrium uH2A. Up to date, the degradation of uH2A was shown to be catalyzed by uH2A lyase, however no enzymes (factors) concerning its synthesis have been elucidated. Here, we show that ATP is prerequisite for the synthesis of uH2A, and that the disappearance of uH2A at the non-permissive temperature may be due to a reduction in the rate of synthesis rather than an increase in the rate of its degradation.
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PMID:Decrease in uH2A (protein A24) of a mouse temperature-sensitive mutant. 629 86

The nuclear translocation of NF-kappa B follows the degradation of its inhibitor, I kappa B alpha, an event coupled with stimulation-dependent inhibitor phosphorylation. Prevention of the stimulation-dependent phosphorylation of I kappa B alpha, either by treating cells with various reagents or by mutagenesis of certain putative I kappa B alpha phosphorylation sites, abolishes the inducible degradation of I kappa B alpha. Yet, the mechanism coupling the stimulation-induced phosphorylation with the degradation has not been resolved. Recent reports suggest a role for the proteasome in I kappa B alpha degradation, but the mode of substrate recognition and the involvement of ubiquitin conjugation as a targeting signal have not been addressed. We show that of the two forms of I kappa B alpha recovered from stimulated cells in a complex with RelA and p50, only the newly phosphorylated form, pI kappa B alpha, is a substrate for an in vitro reconstituted ubiquitin-proteasome system. Proteolysis requires ATP, ubiquitin, a specific ubiquitin-conjugating enzyme, and other ubiquitin-proteasome components. In vivo, inducible I kappa B alpha degradation requires a functional ubiquitin-activating enzyme and is associated with the appearance of high molecular weight adducts of I kappa B alpha. Ubiquitin-mediated protein degradation may, therefore, constitute an integral step of a signal transduction process.
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PMID:Stimulation-dependent I kappa B alpha phosphorylation marks the NF-kappa B inhibitor for degradation via the ubiquitin-proteasome pathway. 747 48

Ubiquitinated proteins are components of intraneuronal inclusions found in several degenerative diseases. Immunohistochemical studies of neurofilament accumulations in Lewy bodies suggest their possible ubiquitination. We investigated in the present work the presence and the nature of ubiquitin epitopes in purified neurofilament preparations from spinal cord. Ubiquitin antibodies consistently label the medium molecular weight neurofilament subunit, and to a lower extent the two other subunits of the neurofilament triplet. Ubiquitinated neurofilament epitopes are removed in vitro by incubation of neurofilaments with a deubiquitinase purified from nervous tissues. Studies of neurofilament degradation in vitro revealed that addition of ATP and exogenous ubiquitin stimulates the proteolysis of neurofilament by crude soluble fractions from nervous tissues. These observations favor the hypothesis of a physiological function of ubiquitin-associated pathways in degradation of neurofilaments in situ.
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PMID:Possible involvement of ubiquitination in neurofilament degradation. 750 32

Ubiquitin (Ub) activation by the Ub-activating (E1) enzyme is the initial and essential step common to all of the known processes that involve post-translational conjugation of Ub to itself or other proteins. The "activated" Ub, linked via a thioester bond to a specific cysteine residue in one of several Ub-conjugating (E2) enzymes, which catalyze the formation of isopeptide bonds between the C-terminal glycine of Ub and lysine residues of acceptor proteins. In the yeast Saccharomyces cerevisiae, a 114-kDa E1 enzyme is encoded by an essential gene termed UBA1 (McGrath, J.P., Jentsch, S., and Varshavsky, A. (1991) EMBO J. 10, 227-236). We describe the isolation and analysis of another essential gene, termed UBA2, that encodes a 71-kDa protein with extensive sequence similarities to both the UBA1-encoded yeast E1 and E1 enzymes of other organisms. The regions of similarities between Uba1p and Uba2p encompass a putative ATP-binding site as well as a sequence that is highly conserved between the known E1 enzymes and contains the active-site cysteine of E1. This cysteine is shown to be required for an essential function of Uba2p, suggesting that Uba2p-catalyzed reactions involved a transient thioester bond between Uba2p and either Ub or another protein. Uba2p is located largely in the nucleus. The putative nuclear localization signal of Uba2p is near its C terminus. The Uba1p (E1 enzyme) and Uba2p cannot complement each others essential functions even if their subcellular localization is altered by mutagenesis. Uba2p appears to interact with itself and several other S. cerevisiae proteins with apparent molecular masses of 52, 63, 87, and 120 kDa. Uba2p is multiubiquitinated in vivo, suggesting that at least a fraction of Uba2p is metabolically unstable. Uba2p is likely to be a component of the Ub system that functions as either an E2 or E1/E2 enzyme.
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PMID:An essential yeast gene encoding a homolog of ubiquitin-activating enzyme. 762 21

Ubiquitin-125I-alpha-globin conjugate fractions containing either one (Ub1-alpha), or two (Ub2-alpha), or a mixture of three and four (Ub3,4-alpha) molecules of ubiquitin (Ub), covalently linked to one 125I-alpha-globin molecule were isolated after incubation of a proteolysis reaction mixture containing ATP, ubiquitin aldehyde-treated reticulocyte lysate, and human 125I-alpha-globin. Each of the purified conjugate fractions or an identically-purified control sample of unconjugated 125I-alpha-globin was incubated as a substrate in companion proteolysis reaction mixtures containing either purified 26S or 20S rabbit reticulocyte proteasomes. The initial rate of ATP-dependent degradation of the Ub1-alpha conjugate by the 26S proteasomes was approximately 0.44% (1.1 fmol)/min while that of the free 125I-alpha-globin was undetectable. The initial rates of ATP-dependent degradation by the 26S proteasomes of the Ub2-alpha and Ub3,4-alpha conjugates were 2- to-3-fold that of the Ub1-alpha species. Conversely, the degradation of free 125I-alpha-globin and its ubiquitinated conjugates by the 20S proteasomes was not dependent on ATP, nor did it increase with the size of the Ub adduct. Analysis of the products of a reaction mixture with 26S proteasomes by sodium dodecyl sulfate-polyacrylamide gel electrophoresis showed no conversion of the Ub1-alpha conjugate substrate to higher-molecular-mass conjugates. These results suggest that monobiquitinated alpha-globin can be degraded significantly and specifically by interaction directly with the 26S proteasomes.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Degradation of monoubiquitinated alpha-globin by 26S proteasomes. 769 67

Ubiquitination of red blood cell (RBC) proteins was investigated by encapsulation of 125I-ubiquitin into human erythrocytes using a procedure of hypotonic dialysis, isotonic resealing, and reannealing. Incubation (37 degrees C, up to 2 h) of 125I-ubiquitin-loaded cells resulted in the recovery of 125I-ubiquitin with the cytosolic proteins (9.22 +/- 0.4 micrograms/ml RBC) and conjugated to membrane proteins (2.18 +/- 0.05 micrograms/ml RBC). This conjugation was time-dependent, and the predominant membrane protein band that became labeled showed an apparent molecular mass of 240 kDa on SDS-polyacrylamide gel electrophoresis (PAGE). Western blotting experiments with three different anti-ubiquitin antibodies revealed that this protein is also ubiquitinated in vivo. Cell-free experiments have shown that fraction II (a DEAE-bound protein fraction eluted by 0.5 M KCl) prepared from both mature erythrocytes and reticulocytes is able to conjugate ubiquitin to this protein. Ubiquitin conjugation was ATP-dependent (Km 0.09 mM), time-dependent, and fraction II-dependent (8 +/- 0.5 pmol of 125I-ubiquitin/h/mg of fraction II). Isolation of the major RBC membrane protein that is ubiquitinated was obtained by using biotinylated ubiquitin. Membrane proteins, once ubiquitinated with this derivative, were extracted and purified by affinity chromatography on immobilized avidin. The major components retained by the column were two peptides of molecular masses 220 and 240 kDa. Both proteins are recognized by a monoclonal anti-spectrin antibody, but only the 240-kDa component is detected by streptavidin peroxidase conjugate. That indeed the ubiquitinated membrane protein of 240-kDa is alpha-spectrin was confirmed by immunoaffinity chromatography using 125I-ubiquitin and a monoclonal anti-spectrin antibody. Antigen-antibody complexes were purified by protein A chromatography and analyzed by SDS-PAGE and autoradiography. Again two bands of 240 and 220 kDa were eluted (alpha- and beta-spectrin), but only one band corresponding to the electrophoretic mobility of alpha-spectrin was detected by autoradiography. Thus, alpha-spectrin is a substrate for the ATP-dependent ubiquitination system, suggesting that the cytoskeleton is covalently modified by ubiquitination both in reticulocytes and mature RBC.
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PMID:Ubiquitin is conjugated to the cytoskeletal protein alpha-spectrin in mature erythrocytes. 772 1

Most of the increased protein degradation in muscle atrophy caused by starvation and denervation is due to activation of a non-lysosomal ATP-dependent proteolytic process. To determine whether expression of the ubiquitin-proteasome-dependent pathway is activated in atrophying muscles, we measured the levels of mRNA for ubiquitin (Ub) and proteasome subunits, and Ub content. After rats had been deprived of food for 1 or 2 days, the concentration of the two polyubiquitin (polyUb) transcripts increased 2-4-fold in the pale extensor digitorum longus muscle and 1-2.5-fold in the red soleus, whereas total muscle RNA and total mRNA content fell by 50%. After denervation of the soleus, there was a progressive 2-3-fold increase in polyUb mRNA for 1-3 days, whereas total RNA content fell. On starvation or denervation, Ub concentration in the muscles also rose by 60-90%. During starvation, polyUb mRNA levels also increased in heart, but not in liver, kidney, spleen, fat, brain or testes. Although the polyUb gene is a heat-shock gene that is induced in muscles under certain stressful conditions, the muscles of starving rats or after denervation did not express other heat-shock genes. On starvation or denervation, mRNA for several proteasome subunits (C-1, C-3, C-5, C-8 and C-9) also increased 2-4-fold in the atrophying muscles. When the food-deprived animals were re-fed, levels of Ub and proteasome mRNA in their muscles returned to control values within 1 day. In contrast, no change occurred in the levels of muscle mRNAs encoding cathepsin L, cathepsin D and calpain 1 on denervation or food deprivation. Thus polyUb and proteasome mRNAs increased in atrophying muscles in co-ordination with activation of the ATP-dependent proteolytic process.
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PMID:Increase in levels of polyubiquitin and proteasome mRNA in skeletal muscle during starvation and denervation atrophy. 774 90

Ubiquitin modification of many protein targets within cells plays important roles in a variety of biological processes. Among these are regulation of gene expression, regulation of cell cycle and division, involvement in the cellular stress response, modification of cell surface receptors, DNA repair, import of proteins into mitochondria, uptake of precursors of neurotransmitters into synaptosomes, biogenesis of peroxisomes, assembly of ribosomes, and programmed cell death. The mechanisms that underlie these complex processes are poorly understood. The best studied modification occurs in the ubiquitin-mediated proteolytic pathway. Recent experimental evidence indicates that the ubiquitin system is involved in the degradation of mitotic cyclins, oncoproteins and tumor suppressors, in the removal of abnormal and otherwise damaged proteins, and in processing of antigens restricted to class I MHC molecules. Degradation of a protein via the ubiquitin system involves two discrete steps. Initially, multiple ubiquitin molecules are covalently linked in an ATP-dependent mode to the protein substrate. The targeted protein is then degraded by a specific, energy-dependent and high molecular mass protease complex into peptides and free amino acids, and free and reutilizable ubiquitin is released. In addition, stable mono-ubiquitin adducts are also found in the cell, for example, those involving nucleosomal histones. Despite the considerable progress that has been made in elucidating the mode of action and roles of the ubiquitin system, many problems remain unsolved. For example, little is known on the signals that target proteins for degradation. While a few proteins are targeted for degradation following recognition of their N-terminal amino acid residue, the vast majority of cellular proteins are targeted by other signals. The identity of the native cellular substrates of the system is another important, yet unresolved problem: only a few proteins have been recognized so far as substrates of the system in vivo. The scope of this review is to discuss the mechanisms involved in ubiquitin activation, selection of substrates for conjugation, and degradation of ubiquitin-conjugated proteins in the cell-free system. In addition, we shall summarize what is currently known of the physiological roles of ubiquitin-mediated proteolysis in vivo.
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PMID:The ubiquitin-mediated proteolytic pathway: mechanisms of action and cellular physiology. 784 Aug 98

The encephalomyocarditis virus 3C protease has been shown to be rapidly degraded in infected cells and in vitro in rabbit reticulocyte lysate. The in vitro degradation, at least, is accomplished by a virus-independent, ATP-dependent proteolytic system. Here we identify this proteolytic system as the ubiquitin-mediated system. Incubation of the 3C protease in rabbit reticulocyte or cultured mouse cell lysate preparations, alone or in the presence of added ubiquitin or methylated ubiquitin, resulted in the generation of new higher molecular weight species. These new products were shown to be 3C protease-ubiquitin conjugates by their ability to bind antibodies against both the 3C protease and ubiquitin. Supplemental ubiquitin also stimulated the degradation of the 3C protease in these preparations. Large 3C protease-polyubiquitin conjugates were observed to accumulate in reticulocyte lysate in the presence of adenosine 5'-O-(3-thiotriphosphate), an inhibitor of the 26 S multicatalytic protease. This, combined with the fact that the proteolytic activity could be removed from the lysate by sedimentation, implicates the multicatalytic protease in the degradation of the 3C protease-ubiquitin conjugates. It was also found that the slow rate of degradation of a model polyprotein, which resembles the stable viral 3CD diprotein produced in vivo, is likely due to the fact that the polyprotein is a poor substrate for the ubiquitin-conjugating system.
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PMID:The encephalomyocarditis virus 3C protease is a substrate for the ubiquitin-mediated proteolytic system. 796 84


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