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Thermodynamic Examination of the Bulged‐G Motif in the 23S Ribosomal RNA
Author(s) -
Aman Zachary
Publication year - 2017
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.31.1_supplement.599.5
Subject(s) - isothermal titration calorimetry , chemistry , rna , crystallography , nucleotide , ribosome , magnesium , phosphodiester bond , base pair , stereochemistry , biochemistry , dna , organic chemistry , gene
Ribosomal RNA contains several bulged‐G motifs (also known as sarcin‐ricin loop, loop E or S‐turn) that are formed by six conserved nucleotides, which form an S‐shape in the phosphodiester backbone. The motif provides a characteristic hydration pattern and geometry that is recognized by proteins that bind to it. The 5′ and 3′ nucleotides of the bulged G serve different functions and the two bend angles of the S vary between 20° to 39° (5′ side of the bulged G) and between 11° to 29° (3′ side of bulged G). In the bulged‐G motif derived from the sarcin‐ricin loop of the 23S subunit of the ribosome, the bulged G nucleotide forms a base triple with the U‐A base pair just above it. The base triple is a proposed site for divalent magnesium ion binding. In this study, we are examining the stability of the bulged‐G motif in the presence and absence of magnesium ions. Several modified constructs are also being examined. Thermal denaturation and isothermal titration calorimetry experiments were performed in 1M KCl and 10 mM MgCl 2 buffers between pH 5.5 and 7. ΔG° 37 of the RNA wild type (WT) construct was −11.2 kcal/mol in potassium ions and increased to −14.0 kcal/mol in 10 mM magnesium ions at pH 7. The WT RNA's interactions with magnesium ions were also studied using isothermal calorimetry and two binding constants, 0.2 and 1 mM were observed in our initial experiments. Modified RNA constructs showed differences in magnesium binding affinity and will be presented. The WT DNA construct showed no differences in stability in 1 M KCl and 10 mM MgCl 2 .