{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,2,7]],"date-time":"2026-02-07T22:21:44Z","timestamp":1770502904744,"version":"3.49.0"},"reference-count":20,"publisher":"Wiley","issue":"2","license":[{"start":{"date-parts":[[2001,10,18]],"date-time":"2001-10-18T00:00:00Z","timestamp":1003363200000},"content-version":"vor","delay-in-days":2656,"URL":"http:\/\/onlinelibrary.wiley.com\/termsAndConditions#vor"}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["FEBS Letters"],"published-print":{"date-parts":[[1994,7,11]]},"abstract":"<jats:p>The wild\u2010type tumor suppressor protein p53 is a short\u2010lived protein that plays important roles in regulation of cell cycle, differentiation, and survival. Mutations that inactivate or alter the tumor suppressor activity of the protein seem to be the most common genetic change in human cancer and are frequently associated with changes in its stability. The ubiquitin system has been implicated in the degradation of p53 both in vivo and in vitro. A mutant cell line that harbors a thermolabile ubiquitin\u2010activating enzyme, E1, fails to degrade p53 at the nonpermissive temperature. Studies in cell\u2010free extracts have shown that covalent attachment of ubiquitin to the protein requires the three conjugating enzymes: E1, a novel species of ubiquitin\u2010carrier protein (ubiquitin\u2010conjugating enzyme; UBC),E2\u2010F1, and an ubiquitin\u2010protein ligase, E3. Recognition of p53 by the ligase is facilitated by formation of a complex between the protein and the human papillomavirus (HPV) oncoprotein E6. Therefore, the ligase has been designated E6\u2010associated protein (E6\u2010AP). However, these in vitro studies have not demonstrated that the conjugates serve as essential intermediates in the proteolytic process. In fact, in many cases, conjugation of ubiquitin to the target protein does not signal its degradation. Thus, it is essential to demonstrate that p53\u2010ubiquitin adducts serve as essential proteolytic intermediates and are recognized and degraded by the 26S protease complex, the proteolytic arm of the ubiquitin pathway. In this study, we demonstrate that conjugates of p53 generated in the presence of purified, E1, E2, E6\u2010AP, E6, ubiquitin and ATP, are specifically recognized by the 26S protease complex and degraded. In contrast, unconjugated p53 remains stable. The ability to reconstitute the system from purified components will enable detailed analysis of the recognition process and the structural motifs involved in targeting the protein for degradation.<\/jats:p>","DOI":"10.1016\/0014-5793(94)00582-6","type":"journal-article","created":{"date-parts":[[2002,7,25]],"date-time":"2002-07-25T07:40:55Z","timestamp":1027582855000},"page":"126-130","source":"Crossref","is-referenced-by-count":35,"title":["Complete reconstitution of conjugation and subsequent degradation of the tumor suppressor protein p53 by purified components of the ubiquitin proteolytic system"],"prefix":"10.1002","volume":"348","author":[{"given":"Dganit","family":"Shkedy","sequence":"first","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Hedva","family":"Gonen","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Beatrice","family":"Bercovich","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Aaron","family":"Ciechanover","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"311","published-online":{"date-parts":[[2001,10,18]]},"reference":[{"key":"e_1_2_1_2_1","doi-asserted-by":"publisher","DOI":"10.1126\/science.2876518"},{"key":"e_1_2_1_3_1","doi-asserted-by":"publisher","DOI":"10.1128\/MCB.14.3.1997"},{"key":"e_1_2_1_4_1","doi-asserted-by":"publisher","DOI":"10.1073\/pnas.88.1.139"},{"key":"e_1_2_1_5_1","doi-asserted-by":"publisher","DOI":"10.1126\/science.2157286"},{"key":"e_1_2_1_6_1","doi-asserted-by":"publisher","DOI":"10.1016\/0092-8674(90)90409-8"},{"key":"e_1_2_1_7_1","doi-asserted-by":"publisher","DOI":"10.1002\/j.1460-2075.1992.tb05487.x"},{"key":"e_1_2_1_8_1","doi-asserted-by":"publisher","DOI":"10.1002\/j.1460-2075.1992.tb05307.x"},{"key":"e_1_2_1_9_1","doi-asserted-by":"publisher","DOI":"10.1038\/346287a0"},{"key":"e_1_2_1_10_1","doi-asserted-by":"publisher","DOI":"10.1016\/S0021-9258(17)36921-1"},{"key":"e_1_2_1_11_1","doi-asserted-by":"publisher","DOI":"10.1016\/S0021-9258(17)36920-X"},{"key":"e_1_2_1_12_1","doi-asserted-by":"publisher","DOI":"10.1016\/0092-8674(93)90384-3"},{"key":"e_1_2_1_13_1","doi-asserted-by":"publisher","DOI":"10.1146\/annurev.bi.61.070192.003553"},{"key":"e_1_2_1_14_1","doi-asserted-by":"publisher","DOI":"10.1128\/MCB.13.8.4918"},{"key":"e_1_2_1_15_1","unstructured":"1989 Cold Spring Harbor Laboratory Press Cold Spring Harbor NY J. 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