3K0J

Crystal structure of the E. coli ThiM riboswitch in complex with thiamine pyrophosphate and the U1A crystallization module

Summary for 3K0J

• Entry DOI: 10.2210/pdb3k0j/pdb
• Descriptor: U1 small nuclear ribonucleoprotein A, RNA (87-MER), THIAMINE DIPHOSPHATE, ... (4 entities in total)
• Functional Keywords: riboswitch, rna, thi-box, thim, u1a protein, acetylation, mrna processing, mrna splicing, nucleus, phosphoprotein, ribonucleoprotein, rna-binding, spliceosome, rna-rna binding protein complex, rna/rna binding protein
• Biological source: Homo sapiens (human)
• Cellular location: Nucleus: P09012
• Total number of polymer chains: 6
• Total formula weight: 101404.16

Authors

Kulshina, N.,Edwards, T.E.,Ferre-D'Amare, A.R. (deposition date: 2009-09-24, release date: 2009-12-22, Last modification date: 2024-02-21)

Primary citation

Kulshina, N.,Edwards, T.E.,Ferre-D'Amare, A.R.
Thermodynamic analysis of ligand binding and ligand binding-induced tertiary structure formation by the thiamine pyrophosphate riboswitch.
Rna, 16:186-196, 2010

PubMed Abstract: The thi-box riboswitch regulates gene expression in response to the intracellular concentration of thiamine pyrophosphate (TPP) in archaea, bacteria, and eukarya. To complement previous biochemical, genetic, and structural studies of this phylogenetically widespread RNA domain, we have characterized its interaction with TPP by isothermal titration calorimetry. This shows that TPP binding is highly dependent on Mg(2+) concentration. The dissociation constant decreases from approximately 200 nM at 0.5 mM Mg(2+) concentration to approximately 9 nM at 2.5 mM Mg(2+) concentration. Binding is enthalpically driven, but the unfavorable entropy of binding decreases as Mg(2+) concentration rises, suggesting that divalent cations serve to pre-organize the RNA. Mutagenesis, biochemical analysis, and a new crystal structure of the riboswitch suggest that a critical element that participates in organizing the riboswitch structure is the tertiary interaction formed between the P3 and L5 regions. This tertiary contact is distant from the TPP binding site, but calorimetric analysis reveals that even subtle mutations in L5 can have readily detectable effects on TPP binding. The thermodynamic signatures of these mutations, namely decreased favorable enthalpy of binding and small effects on entropy of binding, are consistent with the P3-L5 association contributing allosterically to TPP-induced compaction of the RNA.

PubMed: 19948769
DOI: 10.1261/rna.1847310

Experimental method

X-RAY DIFFRACTION (3.1 Å)