Expression, purification, crystallization and preliminary phasing of the heteromerization domain of the tRNA-export and aminoacylation cofactor Arc1p from yeast.
Eukaryotic aminoacyl-tRNA synthetases (aaRSs) must be integrated into an efficient tRNA-export and shuttling machinery. This is reflected by the presence of additional protein-protein interaction domains and a correspondingly higher degree of complex formation in eukaryotic aaRSs. However, the structural basis of interaction between eukaryotic aaRSs and associated protein cofactors has remained elusive. The N-terminal heteromerization domain of the tRNA aminoacylation and export cofactor Arc1p has been cloned from yeast, expressed and purified. Crystals have been obtained belonging to space group C2, with unit-cell parameters a = 222.32, b = 89.46, c = 126.79 angstroms, beta = 99.39 degrees. Calculated Matthews coefficients are compatible with the presence of 10-25 monomers in the asymmetric unit. A complete multiple-wavelength anomalous dispersion data set has been collected from a selenomethionine-substituted crystal at 2.8 angstroms resolution. Preliminary phasing reveals the presence of 20 monomers organized in five tetramers per asymmetric unit.
Simader
Hannes
H
Structural and Computational Research Program, European Molecular Biology Laboratory (EMBL), Meyerhofstrasse 1, D-69117 Heidelberg, Germany.
Suck
Dietrich
D
D000604:Amino Acyl-tRNA Synthetases / Q000378:metabolism; D001692:Biological Transport; D018360:Crystallography, X-Ray; D046911:Macromolecular Substances; D008958:Models, Molecular; D011487:Protein Conformation; D012331:RNA, Fungal / Q000378:metabolism*; D012343:RNA, Transfer / Q000378:metabolism*; D016601:RNA-Binding Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*
C102145:ARC1 protein, S cerevisiae; D046911:Macromolecular Substances; D012331:RNA, Fungal; D016601:RNA-Binding Proteins; D029701:Saccharomyces cerevisiae Proteins; D012343:RNA, Transfer; D000604:Amino Acyl-tRNA Synthetases