title: Characterization, crystallization and preliminary X-ray crystallographic analysis of the Uba5 fragment necessary for high-efficiency activation of Ufm1.
abstract: Uba5 is the smallest ubiquitin-like molecule-activating enzyme and contains an adenylation domain and a C-terminal region. This enzyme only exists in multicellular organisms. The mechanism through which the enzyme recognizes and activates ubiquitin-fold modifier 1 (Ufm1) remains unknown. In this study, Uba5 adenylation domains with different C-terminal region lengths were cloned, expressed and purified. The results of an in vitro truncation assay suggest that Uba5 residues 57-363 comprise the minimal fragment required for the high-efficiency activation of Ufm1. Crystallization of Uba5 residues 57-363 was performed at 277 K using PEG 3350 as the precipitant, and crystals optimized by microseeding diffracted to 2.95 Å resolution, with unit-cell parameters a=b=97.66, c=144.83 Å, α=β=90, γ=120°. There is one molecule in the asymmetric unit; the Matthews coefficient and the solvent content were calculated to be 2.93 Å3 Da(-1) and 58.1%, respectively.
authors.affiliation: MOE Key Laboratory of Protein Science, School of Life Sciences, Tsinghua University, Beijing 100084, People's Republic of China.
authors.forename: Shutao
authors.initials: S
authors.lastname: Xie
mesh_terms: D001483:Base Sequence; D003460:Crystallization; D018360:Crystallography, X-Ray / Q000379:methods*; D017931:DNA Primers; D006801:Humans; D011506:Proteins / Q000737:chemistry*; D044764:Ubiquitin-Activating Enzymes / Q000737:chemistry*
chemical_list: D017931:DNA Primers; D011506:Proteins; C502095:UBA5 protein, human; C502094:UFM1 protein, human; D044764:Ubiquitin-Activating Enzymes
keywords: Uba5; adenylation domain