2012 Fiscal Year Final Research Report
Characterization of deubiquitylating enzymes in yeast
| Project/Area Number |
22687009
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| Research Category |
Grant-in-Aid for Young Scientists (A)
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| Allocation Type | Single-year Grants |
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| Research Field |
Functional biochemistry
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| Research Institution | 公益財団法人東京都医学総合研究所 (2012)
Tokyo Metropolitan Organization for Medical Research (2010-2011) |
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Principal Investigator |
YASUSHI Saeki 公益財団法人東京都医学総合研究所, 生体分子先端研究分野, 主席研究員 (80462779)
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| Project Period (FY) |
2010 – 2012
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| Keywords | 細胞内タンパク質分解 / ユビキチン |
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| Research Abstract |
Protein ubiquitylation is an essential post-translational modification responsible for targeted protein degradation, cellular signaling activation, DNA damage response, and protein trafficking. Deubiquitylating enzymes (DUBs) functions as an eraser of the ubiquitin signals by removing ubiquitin(s) from the ubiquitylated proteins. Despite of their importance, biological significance of DUBs poorly understood. To test enzymatic properties of DUBs, we expressed and purified yeast 20 DUBs by a wheat germ expression system. Although most DUBs did not have chain-type specificities, we found that four DUBs can cleave a ubiquitin precursor Ubi4. Among the proteasomal DUBs, consistently with previous studies, Rpn11 is essential for degradation of the ubiquitinated substrate whereas Ubp6 has an inhibitory effect for the substrate degradation in vitro. However, both DUBs can cleave both K48- and K63-linked ubiquitin chains, suggesting that other factors might specify the proteasomal degradation signal. We also found that Ubp6 facilitates proteasomal assembly by clearing ubiquitylated substrates from assembly precursors by its deubiquitylating activity. Furthermore, we applied parallel reaction monitoring (PRM) method to quantify the ubiquitin chains and analyzed DUB mutants.
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Research Products
(50 results)
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[Journal Article] Structural basis for specific recognition of Rpt1p, an ATPase subunit of 26 S proteasome, by proteasome-dedi-cated chaperone Hsm3p2012
Author(s)
Takagi, K., Kim, S., Yukii, H., Ueno, M., Morishita, R., Endo, Y., Kato, K., Tanaka, K., Saeki, Y. *, and Mizushima, T *, *Correspondences.
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Journal Title
J. Biol. Chem
Volume: 287
Pages: 12172-12182
Peer Reviewed
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[Journal Article] Hyrtioreticulins A-E, indole alkaloids inhibiting the ubiquitin-activating enzyme, from the marine sponge Hyrtios reticulatus2012
Author(s)
Yamanokuchi, R., Imada, K., Miyazaki, M., Kato, H., Watanabe, T., Fujimuro, M., Saeki, Y., Yoshinaga, S., Terasawa, H., Iwasaki, N., Rotinsulu, H., Losung, F., Mangindaan, R.E., Namikoshi, M., de Voogd, N.J., Yokosawa, H., Tsukamoto, S
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Journal Title
Bioorg. Med. Chem
Volume: 20
Pages: 4437-4442
Peer Reviewed
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[Journal Article] New crystal structure of the proteasome-dedicated chaperone Rpn14 at 1.6 A resolution. Acta. Crystallogr2012
Author(s)
Kim, S., Nishide, A., Saeki, Y., Takagi, K., Tanaka, K., Kato, K., Mizushima, T
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Journal Title
Sect. F. Struct. Biol. Cryst. Commun
Volume: 68
Pages: 517-521
Peer Reviewed
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[Journal Article] Insulin/insulin-like growth factor (IGF) stimulation abrogates an association between a deubiquitinating enzyme USP7 and insulin receptor substrates (IRSs) followed by proteasomal degradation of IRSs2012
Author(s)
Yoshihara, H., Fukushima T, Hakuno, F., Saeki, Y., Tanaka, K., Ito, A., Yoshida, M., Iemura, S., Natsume, T., Asano, T., Chida, K., Girnita, L., Takahashi, S
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Journal Title
Biochem. Biophys. Res. Commun
Volume: 423
Pages: 122-127
Peer Reviewed
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[Journal Article] Localization of the proteasomal ubiquitin receptors Rpn10 and Rpn13 by electron cryomicroscopy2012
Author(s)
Sakata, E., Bohn, S., Mihalache, O., Kiss, P., Beck, B., Nagy, I., Nickell, S., Tanaka, K., Saeki, Y., Forster, F., and Baumeister, W
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Journal Title
Proc. Natl. Acad. Sci. USA
Volume: 109
Pages: 1479-1484
Peer Reviewed
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[Journal Article] The catalytic activity of Ubp6 enhances maturation of the proteasomal regulatory particle2011
Author(s)
Sakata, E., Stengel, F., Fukunaga, K., Zhou, M., Saeki, Y., Forster, F., Baumeister, W., Tanaka, K., and Robinson, C.V
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Journal Title
Mol Cell
Volume: 42
Pages: 637-649
Peer Reviewed
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[Journal Article] Sharpin is a component of the NF-.B activating linear ubiquitin chain assembly complex2011
Author(s)
Tokunaga, F., Nakagawa, T., Nakahara, M., Saeki, Y., Taniguchi, M., Sakata, S., Tanaka, K., Nakano, H., and Iwai, K
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Journal Title
Nature
Volume: 471
Pages: 633-636
Peer Reviewed
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[Journal Article] Polyubiquitin conjugation to NEMO by tripartite motif protein 23 (TRIM23) is critical in antiviral defence2010
Author(s)
Arimoto, K.I., Funami, K., Saeki, Y., Tanaka, K., Okawa, K., Takeuchi, O., Akira, S., Murakami, Y., and Shimotohno, K
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Journal Title
Proc. Natl. Acad. Sci. USA
Volume: 107
Pages: 15856-15861
Peer Reviewed
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[Journal Article] Crystal structure of yeast Rpn14, a chaperone of the 19S regulatory particle of the proteasome2010
Author(s)
Kim, S. *, Saeki, Y. *, Fukunaga, K., Suzuki, A., Takagi, K., Yamane, T., Tanaka, K., Mizushima, T., and Kato, K, *Equal contribution
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Journal Title
J. Biol. Chem
Volume: 285
Pages: 15159-15166
Peer Reviewed
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[Presentation] The 26S proteasome completes its assembly process in the cytoplasm prior to the nuclear translocation2011
Author(s)
Haruka Yukii, Yasushi Saeki, Changi Pack, Keisuke Fukunaga, Eri Sakata, Yasushi Sako, Akio Toh-e, Wolfgang Baumeister, Keiji Tanaka
Organizer
EMBO Conference, Ubiquitin and ubiquitin-like modifiers
Place of Presentation
Cavtat, Croatia
Year and Date
20110921-25
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