Evidence for bidentate substrate binding as the basis for the K48 linkage specificity of otubain 1.
Evidence for bidentate substrate binding as the basis for the K48 linkage specificity of otubain 1.
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DOI：
10.1016/j.jmb.2008.12.085
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发表时间：
2009-03-06
影响因子：
5.6
通讯作者：
Wolberger, Cynthia
中科院分区：
文献类型：
--
作者：
Wang, Tao;Yin, Luming;Cooper, Eric M.;Lai, Ming-Yih;Dickey, Seth;Pickart, Cecile M.;Fushman, David;Wilkinson, Keith D.;Cohen, Robert E.;Wolberger, Cynthia
Otubain 1属于半胱氨酸蛋白酶去泛素化酶的卵巢肿瘤（OTU）结构域类别。我们发现人otubain 1（hOtu 1）是高度连接特异性的，切割Lys 48（K48）连接的多聚泛素，但不切割K63、K29、K6或K11连接的多聚泛素或线性α连接的多聚泛素。切割不限于聚泛素链的任一端，游离和底物连接的聚泛素都被分解。有趣的是，K48-双泛素被hOtu 1切割可以被各种连接类型的双泛素以及单泛素抑制。NMR研究和活性测定表明，近端和远端单位的K48-双泛素结合hOtu 1。Cys 23与泛素-乙烯砜的反应鉴定了与包括Cys 91的活性位点不同的泛素结合位点。需要占据活性位点以能够与第二位点紧密结合。我们建议，不同的结合位点的可切割的债券的两侧的泛素允许hOtu 1区分不同的异肽连接在聚泛素底物。双齿结合可能是一种用于实现连接特异性去泛素化的通用策略。
Otubain 1 belongs to the ovarian tumor (OTU) domain class of cysteine protease deubiquitinating enzymes. We show here that human otubain 1 (hOtu1) is highly linkage-specific, cleaving Lys48 (K48)-linked polyubiquitin but not K63-, K29-, K6-, or K11-linked polyubiquitin, or linear α-linked polyubiquitin. Cleavage is not limited to either end of a polyubiquitin chain, and both free and substrate-linked polyubiquitin are disassembled. Intriguingly, cleavage of K48-diubiquitin by hOtu1 can be inhibited by diubiquitins of various linkage types, as well as by monoubiquitin. NMR studies and activity assays suggest that both the proximal and distal units of K48-diubiquitin bind to hOtu1. Reaction of Cys23 with ubiquitin-vinylsulfone identified a ubiquitin binding site that is distinct from the active site, which includes Cys91. Occupancy of the active site is needed to enable tight binding to the second site. We propose that distinct binding sites for the ubiquitins on either side of the scissile bond allow hOtu1 to discriminate among different isopeptide linkages in polyubiquitin substrates. Bidentate binding may be a general strategy used to achieve linkage-specific deubiquitination.
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影响因子：
4.1
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