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Updated: Jun 22, 2026
Isolation of Translating Ribosomes Containing Peptidyl-tRNAs for Functional and Structural Analyses
Published on: February 25, 2011
Isolation, crystallization and preliminary X-ray analysis of the transamidosome, a ribonucleoprotein involved in
Marc Bailly1, Mickael Blaise, Bernard Lorber
1UPR 9002 'Architecture et Réactivité de l'ARN', Université de Strasbourg, Institut de Biologie Moléculaire et Cellulaire du CNRS, 15 Rue René Descartes, F-67084 Strasbourg CEDEX, France.
Abstract:
Thermus thermophilus deprived of asparagine synthetase synthesizes Asn on tRNA(Asn) via a tRNA-dependent pathway involving a nondiscriminating aspartyl-tRNA synthetase that charges Asp onto tRNA(Asn) prior to conversion of the Asp to Asn by GatCAB, a tRNA-dependent amidotransferase. This pathway also constitutes the route of Asn-tRNA(Asn) formation by bacteria and archaea deprived of asparaginyl-tRNA synthetase. The partners involved in tRNA-dependent Asn formation in T. thermophilus assemble into a ternary complex called the transamidosome. This particule produces Asn-tRNA(Asn) in the presence of free Asp, ATP and an amido-group donor. Crystals of the transamidosome from T. thermophilus were obtained in the presence of PEG 4000 in MES-NaOH buffer pH 6.5. They belonged to the primitive monoclinic space group P2(1), with unit-cell parameters a = 115.9, b = 214.0, c = 127.8 A, beta = 93.3 degrees . A complete data set was collected to 3 A resolution. Here, the isolation and crystallization of the transamidosome from T. thermophilus and preliminary crystallographic data are reported.
