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Profiling the Cross Reactivity of Ubiquitin with the Nedd8 Activating Enzyme by Phage Display

Please always quote using this URN: urn:nbn:de:bvb:20-opus-128479
  • The C-terminal peptides of ubiquitin (UB) and UB-like proteins (UBLs) play a key role in their recognition by the specific activating enzymes (E1s) to launch their transfer through the respective enzymatic cascades thus modifying cellular proteins. UB and Nedd8, a UBL regulating the activity of cullin-RING UB ligases, only differ by one residue at their C-termini; yet each has its specific E1 for the activation reaction. It has been reported recently that UAE can cross react with Nedd8 to enable its passage through the UB transfer cascade forThe C-terminal peptides of ubiquitin (UB) and UB-like proteins (UBLs) play a key role in their recognition by the specific activating enzymes (E1s) to launch their transfer through the respective enzymatic cascades thus modifying cellular proteins. UB and Nedd8, a UBL regulating the activity of cullin-RING UB ligases, only differ by one residue at their C-termini; yet each has its specific E1 for the activation reaction. It has been reported recently that UAE can cross react with Nedd8 to enable its passage through the UB transfer cascade for protein neddylation. To elucidate differences in UB recognition by UAE and NAE, we carried out phage selection of a UB library with randomized C-terminal sequences based on the catalytic formation of UB similar to NAE thioester conjugates. Our results confirmed the previous finding that residue 72 of UB plays a "gate-keeping" role in E1 selectivity. We also found that diverse sequences flanking residue 72 at the UB C-terminus can be accommodated by NAE for activation. Furthermore heptameric peptides derived from the C-terminal sequences of UB variants selected for NAE activation can function as mimics of Nedd8 to form thioester conjugates with NAE and the downstream E2 enzyme Ubc12 in the Nedd8 transfer cascade. Once the peptides are charged onto the cascade enzymes, the full-length Nedd8 protein is effectively blocked from passing through the cascade for the critical modification of cullin. We have thus identified a new class of inhibitors of protein neddylation based on the profiles of the UB C-terminal sequences recognized by NAE.show moreshow less

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Metadaten
Author: Bo Zhao, Keya Zhang, Karan Bhuripanyo, Chan Hee J. Choi, Eric B. Villhauer, Heng Li, Ning Zheng, Hiroaki Kiyokawa, Hermann Schindelin, Jun Yin
URN:urn:nbn:de:bvb:20-opus-128479
Document Type:Journal article
Faculties:Fakultät für Biologie / Rudolf-Virchow-Zentrum
Language:English
Parent Title (English):PLoS ONE
ISSN:1932-6203
Year of Completion:2013
Volume:8
Issue:e70312
Source:PLoS ONE 8(8): e70312. doi:10.1371/journal.pone.0070312
DOI:https://doi.org/10.1371/journal.pone.0070312
Dewey Decimal Classification:6 Technik, Medizin, angewandte Wissenschaften / 61 Medizin und Gesundheit / 610 Medizin und Gesundheit
Tag:E1; complex; conjugation; expression; ligases; neddylation; pathway; protein NEDD8; purification; system
Release Date:2016/05/10
Licence (German):License LogoCC BY: Creative-Commons-Lizenz: Namensnennung