Abstract
Post-translational modification by the small ubiquitin-like modifier (SUMO) is an important mechanism regulating protein function. Identification of SUMO conjugation sites on substrates is a challenging task. Here we employed a proteomic method to map SUMO acceptor lysines in budding yeast proteins. We report the identification of 257 lysine residues where SUMO is potentially attached. Amongst the hits, we identified already known SUMO substrates and sites, confirming the success of the approach. In addition, we tested several of the novel substrates using SUMO immunoprecipitation analysis and confirmed that the SUMO acceptor lysines identified in these proteins are indeed bona fide SUMOylation sites. We believe that the collection of SUMO sites presented here is an important resource for future functional studies of SUMOylation in yeast.
Highlights
small ubiquitin-like modifier (SUMO) is a 10-kDa highly conserved protein modifier that reversibly conjugates to specific lysine residues on many target proteins [1]
The excised gel fragments containing the peptides modified were digested with trypsin, which cleaves after arginine and lysine, and removes most of Smt3-KallR-I96R from the substrate peptides
Identifying lysine residues where SUMO is conjugated to substrates is a challenging process that in many cases is resolved by site-directed mutagenesis of potential acceptor lysines
Summary
SUMO (small ubiquitin-related modifier) is a 10-kDa highly conserved protein modifier that reversibly conjugates to specific lysine residues on many target proteins [1]. The functional consequences of SUMO modification include changes in protein stability, localization, DNA-binding, or protein interactions. These SUMO effects can be mediated by providing a new binding interface, masking existing binding sites, or inducing conformational changes on the substrates [2]. SUMO itself can be further SUMOylated via addition of SUMO moieties to lysine residues on SUMO [3]. SUMO can be released from substrates by SUMO proteases (Ulp and Ulp in yeast)
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