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

1. Ubiquitin has been isolated from bovine erythrocytes by procedures in which the hemoglobin was removed by denaturation with either ethanol-chloroform mixtures or by heating. 2. The proteins soluble to the denaturation step were removed by 3% sodium trichloroacetate (TCA) at pH 2.0-2.5 or by 5% TCA. 3. Ubiquitin was isolated in relatively high yield from the TCA insoluble fraction by use of single ion-exchange chromatographic and gel permeation steps. 4. Ubiquitin shows relatively little cross-linking upon treatment with glutaraldehyde or with dimethyl suberimidate. Heating of the glutaraldehyde treated material in 4 M guanidine, however, leads to marked aggregation. 5. The polymers of ubiquitin react strongly with antibody in an immunoblot assay.
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PMID:Simplified methods for isolation of ubiquitin from erythrocytes. Generation of ubiquitin polymers. 282 33

Ubiquitin was radiolabeled by reaction with 125I-Bolton-Hunter reagent and introduced into HeLa cells using erythrocyte-mediated microinjection. The injected cells were then incubated at 45 degrees C for 5 min (reversible heat-shock) or for 30 min (lethal heat-shock). After either treatment, there were dramatic changes in the levels of ubiquitin conjugates. Under normal culture conditions, approximately 10% of the injected ubiquitin is linked to histones, 40% is found in conjugates with molecular weights greater than 25,000, and the rest is unconjugated. After heat-shock, the free ubiquitin pool and the level of histone-ubiquitin conjugates decreased rapidly, and high molecular weight conjugates predominated. Formation of large conjugates did not require protein synthesis; when analyzed by two-dimensional electrophoresis, the major conjugates did not co-migrate with heat-shock proteins before or after thermal stress. Concomitant with the loss of free ubiquitin, the degradation of endogenous proteins, injected hemoglobin, BSA, and ubiquitin was reduced in heat-shocked HeLa cells. After reversible heat-shock, the decrease in proteolysis was small, and both the rate of proteolysis and the size of the free ubiquitin pool returned to control levels upon incubation at 37 degrees C. In contrast, neither proteolysis nor free ubiquitin pools returned to control levels after lethal heat-shock. However, lethally heat-shocked cells degraded denatured hemoglobin more rapidly than native hemoglobin and ubiquitin-globin conjugates formed within them. Therefore, stabilization of proteins after heat-shock cannot be due to the loss of ubiquitin conjugation or inability to degrade proteins that form conjugates with ubiquitin.
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PMID:Microinjection of ubiquitin: changes in protein degradation in HeLa cells subjected to heat-shock. 302 42

Ubiquitin was radioiodinated and introduced into HeLa cells by the erythrocyte-mediated fusion procedure. Fractionation of injected HeLa cells and subsequent NaDodSO4/polyacrylamide gel electrophoresis showed that HeLa nuclei contained two major labeled proteins: ubiquitin and the histone H2A-ubiquitin conjugate, protein A24. HeLa cytosol contained ubiquitin and a series of ubiquitin-protein conjugates of diverse molecular weights. When injected HeLa cells were treated with phenylhydrazine to denature the cotransferred hemoglobin, a series of prominent ubiquitin-globin conjugates appeared. The identity of these conjugates was established by microinjection experiments in which both proteins were labeled. At low doses of phenylhydrazine, the intracellular concentration of globin-ubiquitin conjugates was proportional to the rate of hemoglobin degradation. This result, together with the observation that ubiquitin conjugation to globin is markedly enhanced by phenylhydrazine-induced denaturation of hemoglobin, provides support for the hypothesis that the covalent attachment of ubiquitin to proteins signals proteolysis.
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PMID:Conjugation of ubiquitin to denatured hemoglobin is proportional to the rate of hemoglobin degradation in HeLa cells. 631 May 49