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.
Structural and Computational Research Program, European Molecular Biology Laboratory (EMBL), Meyerhofstrasse 1, D-69117 Heidelberg, Germany.
Hannes
H
Simader
Dietrich
D
Suck
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