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Structural dynamics of ribosome subunit association studied by mixing-spraying time-resolved cryogenic electron microscopy.
Chen, Bo; Kaledhonkar, Sandip; Sun, Ming; Shen, Bingxin; Lu, Zonghuan; Barnard, David; Lu, Toh-Ming; Gonzalez, Ruben L; Frank, Joachim.
Afiliación
  • Chen B; Department of Biochemistry & Molecular Biophysics, Columbia University, New York, NY 10032, USA.
  • Kaledhonkar S; Department of Biochemistry & Molecular Biophysics, Columbia University, New York, NY 10032, USA.
  • Sun M; Department of Biological Sciences, Columbia University, New York, NY 10027, USA.
  • Shen B; Howard Hughes Medical Institute, Columbia University, New York, NY 10032, USA.
  • Lu Z; Center for Integrated Electronics, Rensselaer Polytechnic Institute, Troy, NY 12180, USA.
  • Barnard D; Wadsworth Center, Albany, New York State Department of Health, Albany, NY 12201, USA.
  • Lu TM; Center for Integrated Electronics, Rensselaer Polytechnic Institute, Troy, NY 12180, USA.
  • Gonzalez RL; Department of Chemistry, Columbia University, New York, NY 10027, USA.
  • Frank J; Department of Biochemistry & Molecular Biophysics, Columbia University, New York, NY 10032, USA; Department of Biological Sciences, Columbia University, New York, NY 10027, USA; Howard Hughes Medical Institute, Columbia University, New York, NY 10032, USA. Electronic address: jf2192@cumc.columbi
Structure ; 23(6): 1097-105, 2015 Jun 02.
Article en En | MEDLINE | ID: mdl-26004440
Ribosomal subunit association is a key checkpoint in translation initiation but its structural dynamics are poorly understood. Here, we used a recently developed mixing-spraying, time-resolved, cryogenic electron microscopy (cryo-EM) method to study ribosomal subunit association in the sub-second time range. We have improved this method and increased the cryo-EM data yield by tenfold. Pre-equilibrium states of the association reaction were captured by reacting the mixture of ribosomal subunits for 60 ms and 140 ms. We also identified three distinct ribosome conformations in the associated ribosomes. The observed proportions of these conformations are the same in these two time points, suggesting that ribosomes equilibrate among the three conformations within less than 60 ms upon formation. Our results demonstrate that the mixing-spraying method can capture multiple states of macromolecules during a sub-second reaction. Other fast processes, such as translation initiation, decoding, and ribosome recycling, are amenable to study with this method.
Asunto(s)

Texto completo: 1 Colección: 01-internacional Base de datos: MEDLINE Asunto principal: Modelos Moleculares / Microscopía por Crioelectrón / Escherichia coli / Subunidades Ribosómicas Tipo de estudio: Prognostic_studies / Risk_factors_studies Idioma: En Revista: Structure Asunto de la revista: BIOLOGIA MOLECULAR / BIOQUIMICA / BIOTECNOLOGIA Año: 2015 Tipo del documento: Article País de afiliación: Estados Unidos Pais de publicación: Estados Unidos

Texto completo: 1 Colección: 01-internacional Base de datos: MEDLINE Asunto principal: Modelos Moleculares / Microscopía por Crioelectrón / Escherichia coli / Subunidades Ribosómicas Tipo de estudio: Prognostic_studies / Risk_factors_studies Idioma: En Revista: Structure Asunto de la revista: BIOLOGIA MOLECULAR / BIOQUIMICA / BIOTECNOLOGIA Año: 2015 Tipo del documento: Article País de afiliación: Estados Unidos Pais de publicación: Estados Unidos