Asymmetric nature of two subunits of RAD18, a RING-type ubiquitin ligase E3, in the human RAD6A–RAD18 ternary complex
作者： Yuji Masuda , Miki Suzuki , Hidehiko Kawai , Fumio Suzuki , Kenji Kamiya
DOI: 10.1093/NAR/GKR805
关键词: Ubiquitin 、 Protein subunit 、 Stereochemistry 、 Ligase activity 、 Protein structure 、 Biochemistry 、 DNA ligase 、 Biology 、 Ubiquitin-conjugating enzyme 、 Ternary complex 、 Ubiquitin ligase
摘要: RAD18, a RING-type ubiquitin ligase (E3) that plays an essential role in post-replication repair, possesses distinct domains named RING, UBZ, SAP and the RAD6-binding domain (R6BD) forms dimer. RAD6, ubiquitin-conjugating enzyme (E2), stably associates with R6BD C-terminal portion. In this study, we established method to distinguish between two subunits of RAD18 by introduction different tags, analyzed mutant complexes. Our results, surprisingly, demonstrate RAD6A form ternary complex, RAD6A-(RAD18)(2) presence only one is sufficient for complex formation activity. Interestingly, activity dimer lacking both R6BDs not restored even large amounts added solution, suggesting requirement precise juxtaposition via interaction R6BD. We further show mutations either RING or SAP, but strongly reduce activity, although inactivation without effect. These results suggest asymmetric nature complex.
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