Tandem UIMs confer Lys48 ubiquitin chain substrate preference to deubiquitinase USP25.
Tandem UIMs confer Lys48 ubiquitin chain substrate preference to deubiquitinase USP25.
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DOI：
10.1038/srep45037
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发表时间：
2017-03-22
影响因子：
4.6
通讯作者：
Komada M
中科院分区：
文献类型：
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作者：
Kawaguchi K;Uo K;Tanaka T;Komada M
泛素特异性蛋白酶（USP）25属于USP去泛素化酶家族，具有两个串联的泛素相互作用基序（UIM），一个约20个氨基酸的α-螺旋延伸结合泛素。然而，UIM在USP 25中的作用仍不清楚。在这里，我们表明，串联UIM区结合Lys 48-，但不是Lys 63-，连接的泛素链，其中两个UIM发挥了关键和合作的作用。纯化的USP 25对Lys 48-比Lys 63-连接的泛素链表现出更高的泛素异肽酶活性。破坏串联UIMs的泛素结合能力的突变导致USP 25的泛素异肽酶活性降低，表明UIMs在发挥USP 25的全部催化活性中的作用。此外，当将Lys 48连接的泛素链的结合偏好转换为Lys 63连接的泛素链的突变引入串联UIM区域时，USP 25突变体分别获得了对Lys 63和Lys 48连接的泛素链的升高和降低的异肽酶活性。这些结果表明，串联UIM对Lys 48连接的泛素链的结合偏好不仅有助于完全催化活性，而且有助于USP 25的泛素链底物偏好，可能是通过选择性地将Lys 48连接的泛素链底物保持在催化核心附近。
Ubiquitin-specific protease (USP) 25, belonging to the USP family of deubiquitinases, harbors two tandem ubiquitin-interacting motifs (UIMs), a ~20-amino-acid α-helical stretch that binds to ubiquitin. However, the role of the UIMs in USP25 remains unclear. Here we show that the tandem UIM region binds to Lys48-, but not Lys63-, linked ubiquitin chains, where the two UIMs played a critical and cooperative role. Purified USP25 exhibited higher ubiquitin isopeptidase activity to Lys48-, than to Lys63-, linked ubiquitin chains. Mutations that disrupted the ubiquitin-binding ability of the tandem UIMs resulted in a reduced ubiquitin isopeptidase activity of USP25, suggesting a role for the UIMs in exerting the full catalytic activity of USP25. Moreover, when mutations that convert the binding preference from Lys48- to Lys63-linked ubiquitin chains were introduced into the tandem UIM region, the USP25 mutants acquired elevated and reduced isopeptidase activity toward Lys63- and Lys48-linked ubiquitin chains, respectively. These results suggested that the binding preference of the tandem UIMs toward Lys48-linked ubiquitin chains contributes not only to the full catalytic activity but also to the ubiquitin chain substrate preference of USP25, possibly by selectively holding the Lys48-linked ubiquitin chain substrates in the proximity of the catalytic core.