Interaction network of the ribosome assembly machinery from a eukaryotic thermophile.

Autor: Baßler J; Biochemistry Center Heidelberg BZH, University of Heidelberg, Heidelberg, 69120, Germany., Ahmed YL; Biochemistry Center Heidelberg BZH, University of Heidelberg, Heidelberg, 69120, Germany., Kallas M; Biochemistry Center Heidelberg BZH, University of Heidelberg, Heidelberg, 69120, Germany., Kornprobst M; Biochemistry Center Heidelberg BZH, University of Heidelberg, Heidelberg, 69120, Germany., Calviño FR; Biochemistry Center Heidelberg BZH, University of Heidelberg, Heidelberg, 69120, Germany., Gnädig M; Biochemistry Center Heidelberg BZH, University of Heidelberg, Heidelberg, 69120, Germany., Thoms M; Biochemistry Center Heidelberg BZH, University of Heidelberg, Heidelberg, 69120, Germany., Stier G; Biochemistry Center Heidelberg BZH, University of Heidelberg, Heidelberg, 69120, Germany., Ismail S; Biochemistry Center Heidelberg BZH, University of Heidelberg, Heidelberg, 69120, Germany., Kharde S; Biochemistry Center Heidelberg BZH, University of Heidelberg, Heidelberg, 69120, Germany., Castillo N; Biochemistry Center Heidelberg BZH, University of Heidelberg, Heidelberg, 69120, Germany., Griesel S; Biochemistry Center Heidelberg BZH, University of Heidelberg, Heidelberg, 69120, Germany., Bastuck S; Biochemistry Center Heidelberg BZH, University of Heidelberg, Heidelberg, 69120, Germany., Bradatsch B; Biochemistry Center Heidelberg BZH, University of Heidelberg, Heidelberg, 69120, Germany., Thomson E; Biochemistry Center Heidelberg BZH, University of Heidelberg, Heidelberg, 69120, Germany., Flemming D; Biochemistry Center Heidelberg BZH, University of Heidelberg, Heidelberg, 69120, Germany., Sinning I; Biochemistry Center Heidelberg BZH, University of Heidelberg, Heidelberg, 69120, Germany., Hurt E; Biochemistry Center Heidelberg BZH, University of Heidelberg, Heidelberg, 69120, Germany.
Jazyk: angličtina
Zdroj: Protein science : a publication of the Protein Society [Protein Sci] 2017 Feb; Vol. 26 (2), pp. 327-342. Date of Electronic Publication: 2017 Jan 14.
DOI: 10.1002/pro.3085
Abstrakt: Ribosome biogenesis in eukaryotic cells is a highly dynamic and complex process innately linked to cell proliferation. The assembly of ribosomes is driven by a myriad of biogenesis factors that shape pre-ribosomal particles by processing and folding the ribosomal RNA and incorporating ribosomal proteins. Biochemical approaches allowed the isolation and characterization of pre-ribosomal particles from Saccharomyces cerevisiae, which lead to a spatiotemporal map of biogenesis intermediates along the path from the nucleolus to the cytoplasm. Here, we cloned almost the entire set (∼180) of ribosome biogenesis factors from the thermophilic fungus Chaetomium thermophilum in order to perform an in-depth analysis of their protein-protein interaction network as well as exploring the suitability of these thermostable proteins for structural studies. First, we performed a systematic screen, testing about 80 factors for crystallization and structure determination. Next, we performed a yeast 2-hybrid analysis and tested about 32,000 binary combinations, which identified more than 1000 protein-protein contacts between the thermophilic ribosome assembly factors. To exemplary verify several of these interactions, we performed biochemical reconstitution with the focus on the interaction network between 90S pre-ribosome factors forming the ctUTP-A and ctUTP-B modules, and the Brix-domain containing assembly factors of the pre-60S subunit. Our work provides a rich resource for biochemical reconstitution and structural analyses of the conserved ribosome assembly machinery from a eukaryotic thermophile.
(© 2017 The Protein Society.)
Databáze: MEDLINE
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