Identification of the enzyme required for activation of the small ubiquitin-like protein SUMO-1
Identification of the enzyme required for activation of the small ubiquitin-like protein SUMO-1
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DOI：
10.1074/jbc.274.15.10618
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发表时间：
1999-04-09
影响因子：
4.8
通讯作者：
Hay, RT
中科院分区：
文献类型：
--
作者：
Desterro, JMP;Rodriguez, MS;Hay, RT
泛素样蛋白SUMO-1与多种蛋白结合，包括RAN GTP酶激活蛋白1(RanGAP1)、I kappa Bα和PML，SUMO-1修饰的蛋白显示出改变的亚细胞靶向性和/或稳定性。我们已经从人细胞中纯化了SUMO-1激活酶，发现它含有38和72 kDa两个亚基，每个亚基的cDNA分离表明它们与泛素激活酶和负责Smt3p和Rub-1p结合的酿酒酵母酶同源。在体外，重组SAE1/SAE2(SUMO-1激活酶)能够催化依赖于ATP的SUMO-1和SAE2之间硫酯键的形成。在SAE1/SAE2、Ubch9和ATP存在的情况下，SUMO-1有效地连接到蛋白底物I kappa Bα上。由于SAE1/SAE2、Ubch9、SUMO-1和I kappa Bα都是均一的重组蛋白，因此似乎在体外与I kappa Bα的SUMO-1结合不需要相当于E3泛素蛋白连接酶的活性。
The ubiquitin-like protein SUMO-1 is conjugated to a variety of proteins including Ran GTPase-activating protein 1 (RanGAP1), I kappa B alpha, and PML, SUMO-1-modified proteins display altered subcellular targeting and/or stability. We have purified the SUMO-1-activating enzyme from human cells and shown that it contains two subunits of 38 and 72 kDa, Isolation of cDNAs for each subunit indicates that they are homologous to ubiquitin-activating enzymes and to the Saccharomyces cerevisiae enzymes responsible for conjugation of Smt3p and Rub-1p, In vitro, recombinant SAE1/SAE2 (SUMO-1-activating enzyme) was capable of catalyzing the ATP-dependent formation of a thioester linkage between SUMO-1 and SAE2. The addition of the SUMO-1-conjugating enzyme Ubch9 resulted in efficient transfer of the thioester-linked SUMO-1 from SAE2 to Ubch9, In the presence of SAE1/SAE2, Ubch9, and ATP, SUMO-1 was efficiently conjugated to the protein substrate I kappa B alpha. As SAE1/SAE2, Ubch9, SUMO-1, and I kappa B alpha are all homogeneous, recombinant proteins, it appears that SUMO-1 conjugation of I kappa B alpha in vitro does not require the equivalent of an E3 ubiquitin protein ligase activity.