Preparation and crystallization of riboswitch-ligand complexes Journal Article

Authors: Pikovskaya, O.; Serganov, A.; Polonskaia, A.; Patel, D. J.
Article Title: Preparation and crystallization of riboswitch-ligand complexes
Abstract: Riboswitches are mRNA regions that regulate the expression of genes in response to various cellular metabolites. These RNA sequences, typically situated in the untranslated regions of mRNAs, possess complex structures that dictate highly specific binding to certain ligands, such as nucleobases, coenzymes, amino acids, and sugars, without protein assistance. Depending on the presence of the ligand, metabolite-binding domains of riboswitches can adopt two alternative conformations, which define the conformations of the adjacent sequences involved in the regulation of gene expression. In order to understand in detail the nature of riboswitch-ligand interactions and the molecular basis of riboswitch-based gene expression control, it is necessary to determine the three-dimensional structures of riboswitch-ligand complexes. This chapter outlines the techniques that are employed to prepare riboswitch-ligand complexes for structure determination using X-ray crystallography. The chapter describes the principles of construct design, in vitro transcription, RNA purification, complex formation, and crystallization screening utilized during the successful crystallization of several riboswitches.
Keywords: genetics; molecular genetics; methodology; chemistry; molecular sequence data; nucleotide sequence; ligand; ligands; base sequence; crystallography, x-ray; conformation; nucleic acid conformation; untranslated rna; rna, untranslated; crystallization; x ray crystallography; regulatory rna sequence; synthesis; regulatory sequences, ribonucleic acid
Journal Title: Methods in Molecular Biology
Volume: 540
ISSN: 1064-3745
Publisher: Humana Press Inc  
Date Published: 2009-01-01
Start Page: 115
End Page: 128
Language: English
DOI: 10.1007/978-1-59745-558-9_9
PUBMED: 19381556
PROVIDER: scopus
Notes: Chapter 9 in "Roboswitches: Methods and Protocols" (ISBN: 978-1-934115-88-6) --- Cited By (since 1996): 1 - Export Date: 30 November 2010 - Source: Scopus
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  1. Dinshaw J Patel
    392 Patel