Permanent URL to this publication: http://dx.doi.org/10.5167/uzh-26506
Imkamp, F; Rosenberger, T; Striebel, F; Keller, P M; Amstutz, B; Sander, P; Weber-Ban, E (2010). Deletion of dop in Mycobacterium smegmatis abolishes pupylation of protein substrates in vivo. Molecular Microbiology, 75(3):744-754.
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Proteasome-bearing bacteria make use of a ubiquitin-like modification pathway to target proteins for proteasomal turnover. In a process termed pupylation, proteasomal substrates are covalently modified with the small protein Pup that serves as a degradation signal. Pup is attached to substrate proteins by action of PafA. Prior to its attachment, Pup needs to undergo deamidation at its C-terminal residue, converting glutamine to glutamate. This step is catalyzed in vitro by Dop. In order to characterize Dop activity in vivo, we generated a dop deletion mutant in Mycobacterium smegmatis. In the Deltadop strain, pupylation is severely impaired and the steady-state levels of two known proteasomal substrates are drastically increased. Pupylation can be re-established by complementing the mutant with either DopWt or a Pup variant carrying a glutamate at its ultimate C-terminal position (PupGGE). Our data show that Pup is deamidated by Dop in vivo and that likely Dop alone is responsible for this activity. Furthermore, we demonstrate that a putative N-terminal ATP binding motif is crucial for catalysis, since a single point mutation (E10A) in this motif abolishes Dop activity both in vivo and in vitro.
|Item Type:||Journal Article, refereed, original work|
|Communities & Collections:||04 Faculty of Medicine > Institute of Medical Microbiology|
|DDC:||570 Life sciences; biology|
610 Medicine & health
|Date:||13 January 2010|
|Deposited On:||20 Jan 2010 10:30|
|Last Modified:||28 Nov 2013 02:26|
|Free access at:||Publisher DOI. An embargo period may apply.|
|Citations:||Web of Science®. Times cited: 21|
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