6dg4 | pdb_00006dg4

Structure of the Chaetomium thermophilum Ulp1-like SUMO protease catalytic domain

spinqualitylabelsall models 6dg4, resolution 1.44Å Show:Asymmetric Unit Biological Assembly Drag the structure with the mouse to rotate   Export Animated Image   Print3D Structural highlights 6dg4 is a 1 chain structure with sequence from Cbs 144.50. Full crystallographic information is available from OCA. For a guided tour on the structure components use FirstGlance. Ligands: GOL, SO4 Gene: CTHT_0004370 (CBS 144.50) Activity: Ulp1 peptidase, with EC number 3.4.22.68 Resources: FirstGlance, OCA, PDBe, RCSB, PDBsum, ProSAT Publication Abstract from PubMed Small ubiquitin-like modifier (SUMO) is commonly used as a protein fusion domain to facilitate expression and purification of recombinant proteins, and a SUMO-specific protease is then used to remove SUMO from these proteins. Although this protease is highly specific, its limited solubility and stability hamper its utility as an in vitro reagent. Here, we report improved SUMO protease enzymes obtained via two approaches. First, we developed a computational method and used it to re-engineer wildtype Ulp1 from Saccharomyces cerevisiae to improve protein solubility. Second, we discovered an improved SUMO protease via genomic mining of the thermophilic fungus Chaetomium thermophilum, as proteins from thermophilic organisms are commonly employed as reagent enzymes. Following expression in Escherichia coli, we found these re-engineered enzymes can be more thermostable and up to twelve times more soluble, all while retaining wildtype-or-better levels of SUMO protease activity. The computational method we developed to design solubility-enhancing substitutions is based on the RosettaScripts application for the macromolecular modeling suite Rosetta, and it's broadly applicable for the improvement of solution properties of other proteins. Moreover, we determined the X-ray crystal structure of a SUMO protease from C. thermophilum to 1.44 A resolution. This structure revealed that this enzyme exhibits structural and functional conservation with the S. cerevisiae SUMO protease, despite exhibiting only 28% sequence identity. In summary, by re-engineering the Ulp1 protease and discovering a SUMO protease from C. thermophilum, we have obtained proteases that are more soluble, more thermostable, and more efficient than the current commercially available Ulp1 enzyme. Discovery and engineering of enhanced SUMO protease enzymes.,Lau YK, Baytshtok V, Howard TA, Fiala BM, Morgan JM, Carter LP, Baker D, Lima CD, Bahl CD J Biol Chem. 2018 Jul 5. pii: RA118.004146. doi: 10.1074/jbc.RA118.004146. PMID:29976752 From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.

Proteopedia Page Contributors and Editors (what is this?)

OCA