Two ZnF-UBP Domains in Isopeptidase T (USP5)
详细信息 查看全文
- 作者：George V. Avvakumov ; John R. Walker ; Sheng Xue ; Abdellah Allali-Hassani ; Abdalin Asinas ; Usha B. Nair ; Xianyang Fang ; Xiaobing Zuo ; Yun-Xing Wang ; Keith D. Wilkinson ; Sirano Dhe-Paganon
- 刊名：Biochemistry
- 出版年：2012
- 出版时间：February 14, 2012
- 年：2012
- 卷：51
- 期：6
- 页码：1188-1198
- 全文大小：599K
- 年卷期：v.51,no.6(February 14, 2012)
- ISSN：1520-4995
文摘
Human ubiquitin-specific cysteine protease 5 (USP5, also known as ISOT and isopeptidase T), an 835-residue multidomain enzyme, recycles ubiquitin by hydrolyzing isopeptide bonds in a variety of unanchored polyubiquitin substrates. Activation of the enzyme鈥檚 hydrolytic activity toward ubiquitin-AMC (7-amino-4-methylcoumarin), a fluorogenic substrate, by the addition of free, unanchored monoubiquitin suggested an allosteric mechanism of activation by the ZnF-UBP domain (residues 163鈥?91), which binds the substrate鈥檚 unanchored diglycine carboxyl tail. By determining the structure of full-length USP5, we discovered the existence of a cryptic ZnF-UBP domain (residues 1鈥?56), which was tightly bound to the catalytic core and was indispensable for catalytic activity. In contrast, the previously characterized ZnF-UBP domain did not contribute directly to the active site; a paucity of interactions suggested flexibility between these two domains consistent with an ability by the enzyme to hydrolyze a variety of different polyubiquitin chain linkages. Deletion of the known ZnF-UBP domain did not significantly affect rate of hydrolysis of ubiquitin-AMC and suggested that it is likely associated mainly with substrate targeting and specificity. Together, our findings show that USP5 uses multiple ZnF-UBP domains for substrate targeting and core catalytic function.