The Lysine 48 and Lysine 63 Ubiquitin Conjugates Are Processed Differently by the 26 S Proteasome
The Lysine 48 and Lysine 63 Ubiquitin Conjugates Are Processed Differently by the 26 S Proteasome
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DOI：
10.1074/jbc.m109.052928
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发表时间：
2009-12-18
影响因子：
4.8
通讯作者：
Liu, Chang-Wei
中科院分区：
文献类型：
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作者：
Jacobson, Andrew D.;Zhang, Nan-Yan;Liu, Chang-Wei
Lys-63泛素链在蛋白酶体降解靶向蛋白中的作用尚不清楚。我们系统地比较了Lys-63泛素链的蛋白酶体处理和典型的蛋白分解信号Lys-48泛素链的处理。对HeLa细胞中泛素链的定量质谱分析表明，Lys-63泛素链的水平对短时蛋白酶体抑制不敏感。此外，26 S蛋白酶体结合组分中的Lys-48/Lys-63比细胞裂解产物中的比率高1.7倍，这可能是因为一些细胞内的Lys-63泛素结合物被Lys-63链特异性结合蛋白隔离。在体外，Lys-48和Lys-63泛素链与26个S蛋白酶体的结合能力相当，而Lys-63链的去泛素化速度是Lys-48链的6倍。此外，Lys-63四泛素结合的UbcH10迅速去泛素化成单泛素形式，而Lys-48四泛素以UbcH10为靶标进行降解。此外，我们发现26个S蛋白酶体的泛素醛和1，10-菲咯啉敏感的去泛素化活性都有助于Lys-48和Lys-63的连锁去泛素化，尽管抑制程度不同。综上所述，我们的发现表明，与Lys-48链相比，细胞内的Lys-63链具有较少的蛋白酶体可及性，而与蛋白酶体结合的Lys-63链更快地去泛素化，这可能导致Lys-63结合物的低效降解。
The role of Lys-63 ubiquitin chains in targeting proteins for proteasomal degradation is still obscure. We systematically compared proteasomal processing of Lys-63 ubiquitin chains with that of the canonical proteolytic signal, Lys-48 ubiquitin chains. Quantitative mass spectrometric analysis of ubiquitin chains in HeLa cells determines that the levels of Lys-63 ubiquitin chains are insensitive to short-time proteasome inhibition. Also, the Lys-48/Lys-63 ratio in the 26 S proteasome-bound fraction is 1.7-fold more than that in the cell lysates, likely because some cellular Lys-63 ubiquitin conjugates are sequestered by Lys-63 chain-specific binding proteins. In vitro, Lys-48 and Lys-63 ubiquitin chains bind the 26 S proteasome comparably, whereas Lys-63 chains are deubiquitinated 6-fold faster than Lys- 48 chains. Also, Lys-63 tetraubiquitin-conjugated UbcH10 is rapidly deubiquitinated into the monoubiquitinated form, whereas Lys- 48 tetraubiquitin targets UbcH10 for degradation. Furthermore, we found that both the ubiquitin aldehyde- and 1,10-phenanthroline- sensitive deubiquitinating activities of the 26 S proteasome contribute to Lys-48- and Lys-63-linkage deubiquitination, albeit the inhibitory extents are different. Together, our findings suggest that compared with Lys-48 chains, cellular Lys-63 chains have less proteasomal accessibility, and proteasome-bound Lys-63 chains are more rapidly deubiquitinated, which could cause inefficient degradation of Lys-63 conjugates.