Structural analysis of the multienzyme aminoacyl-tRNA synthetase complex: a three-domain model based on reversible chemical crosslinking.
A subset of eukaryotic aminoacyl-tRNA synthetases (a-RS) are contained in a multienzyme complex for which little structural detail is known. Three reversible chemical crosslinking reagents have been used to investigate the arrangement of polypeptides within this particle as isolated from rabbit reticulocytes. Identification of the crosslinked protein pairs was accomplished by two-dimensional SDS diagonal gel electrophoresis. Seventeen neighboring protein pairs have been identified. Eight are seen with at least two reagents: K-RS:p38, D-RS:K-RS, R-RS dimer, K-RS dimer, K-RS:Q-RS, E/P-RS:K-RS, E/P-RS:I-RS, and Q-RS with one of the nonsynthetase proteins. Nine more are observed with one reagent: D-RS dimer, R-RS:p43, D-RS:Q-RS, D-RS:M-RS, K-RS:L-RS, I-RS:R-RS, D-RS:E/P-RS, I-RS:Q-RS, I-RS:L-RS. One trimeric association is seen: E/P-RS:I-RS:L-RS. The observed neighboring protein pairs suggest that the polypeptides within the aminoacyl-tRNA synthetase complex are distributed in three structural domains of similar mass. These can be arranged in a U-shaped particle in which each "arm" is considered a domain and the third forms the "base" of the structure. The arms have been termed domain I (D-RS, M-RS, Q-RS) and domain II (K-RS, R-RS), with domain III (E/P-RS, I-RS, L-RS) assigned to the base. The smaller proteins (p38, p43) may bridge the domains. This proposed spatial relationship of these domains, as well as their compositions, are consistent with earlier studies. Thus, this study provides an initial three-dimensional working model of the arrangement of polypeptides within the multienzyme aminoacyl-tRNA synthetase complex.
Norcum
M T
MT
Department of Biochemistry, University of Mississippi Medical Center, Jackson 39216-4505, USA. mnorcum@fiona.umsmed.edu
Warrington
J A
JA
D000604:Amino Acyl-tRNA Synthetases / Q000737:chemistry*; D000818:Animals; D002945:Cisplatin / Q000378:metabolism; D003432:Cross-Linking Reagents / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel / Q000379:methods; D008958:Models, Molecular; D010455:Peptides / Q000737:chemistry; D011487:Protein Conformation; D011817:Rabbits; D012156:Reticulocytes / Q000201:enzymology; D013388:Succinimides / Q000378:metabolism
D003432:Cross-Linking Reagents; D010455:Peptides; D013388:Succinimides; C058151:4-succinimidyloxycarbonyl-alpha-methyl-alpha(2-pyridyldithio)toluene; C028722:ethylene glycolylbis(succinimidyl succinate); D000604:Amino Acyl-tRNA Synthetases; D002945:Cisplatin