Phosphorylation and Methylation of Proteasomal Proteins of the Haloarcheon Haloferax volcanii
Proteasomes are composed of 20S core particles (CPs) of- and-type subunits that associate with regulatory particle AAA ATPases such as the proteasome-activating nucleotidase (PAN) complexes of archaea. In this study, the roles and additional sites of post-translational modification of proteasomes were investigated using the archaeonHaloferax volcaniias a model. Indicative of phosphorylation, phosphatase-sensitive isoforms ofandwere detected by 2-DE immunoblot. To map these and other potential sites of post-translational modification, proteasomes were purified and analyzed by tandem mass spectrometry (MS/MS). Using this approach, several phosphosites were mapped includingThr147,2 Thr13/Ser14 and PAN-A Ser340. Multiple methylation sites were also mapped to, thus, revealing a new type of proteasomal modification. Probing the biological role ofand PAN-A phosphorylation by site-directed mutagenesis revealed dominant negative phenotypes for cell viability and/or pigmentation forvariants including Thr147Ala, Thr158Ala and Ser58Ala. AnH. volcaniiRio1p Ser/Thr kinase homolog was purified and shown to catalyze autophosphorylation and phosphotransfer to. Thevariants in Thr and Ser residues that displayed dominant negative phenotypes were significantly reduced in their ability to accept phosphoryl groups from Rio1p, thus, providing an important link between cell physiology and proteasomal phosphorylation.
- Authors:
-
[1]
; [1]
; [1]
; [1]
; [1]
- Department of Microbiology and Cell Science, University of Florida, Gainesville, FL 32611, USA
- Publication Date:
- Grant/Contract Number:
- FG02-05ER15650
- Type:
- Published Article
- Journal Name:
- Archaea
- Additional Journal Information:
- Journal Volume: 2010; Related Information: CHORUS Timestamp: 2016-08-11 11:11:23; Journal ID: ISSN 1472-3646
- Publisher:
- Hindawi
- Sponsoring Org:
- USDOE
- Country of Publication:
- United States
- Language:
- English
- Subject:
- 59 BASIC BIOLOGICAL SCIENCES
- OSTI Identifier:
- 1198438