Structural energetics and base-pair opening dynamics in sarcin-ricin domain RNA.
The sarcin-ricin domain is a universal element of the RNA from the large ribosomal subunit. The domain is part of the binding site for elongation factors and is specifically cleaved by the toxins alpha-sarcin and ricin. In this work, we have mapped the energetics and dynamics of individual structural motifs in a 29-mer RNA oligomer containing the sarcin-ricin domain. The stability of individual base pairs in the structure was characterized from measurements of the exchange rates of imino protons using nuclear magnetic resonance spectroscopy at 10 degrees C. The measurements also provided the rates of opening and closing for selected base pairs. The results reveal that the structural stabilization free energies in the sarcin-ricin domain are broadly distributed between 2.9 and 10.6 kcal/mol. One of the least stable sites in the structure is the noncanonical G-A base pair located next to the phosphodiester bond that is cleaved by alpha-sarcin. The low stability of this base pair supports the proposal that cleavage by alpha-sarcin occurs by a base flipping mechanism. The opening dynamics of other base pairs is affected by elements of the structure such as the bulged-G motif and its cross-strand stacking. Participation in these motifs increases the lifetimes of the bases in an open, solvent-accessible conformation.
Chen
Congju
C
Department of Chemistry and Molecular Biophysics Program, Wesleyan University, Middletown, Connecticut 06459, USA.
Jiang
Lihong
L
Michalczyk
Ryszard
R
Russu
Irina M
IM
D000818:Animals; D020029:Base Pairing / Q000187:drug effects*; D001483:Base Sequence; D004722:Endoribonucleases / Q000737:chemistry; D005656:Fungal Proteins / Q000737:chemistry; D006147:Guanine / Q000737:chemistry; D007097:Imines / Q000737:chemistry; D019906:Nuclear Magnetic Resonance, Biomolecular; D012339:RNA, Ribosomal, 28S / Q000737:chemistry*; D051381:Rats; D012276:Ricin / Q000737:chemistry; D014498:Uracil / Q000737:chemistry
D005656:Fungal Proteins; D007097:Imines; D012339:RNA, Ribosomal, 28S; C014801:alpha-sarcin; D014498:Uracil; D006147:Guanine; D012276:Ricin; D004722:Endoribonucleases