Perturbation of the hierarchical folding of a large RNA by the destabilization of its Scaffold's tertiary structure.
The P4-P6 domain serves as a scaffold against which the periphery and catalytic core organize and fold during Mg2+-mediated folding of the Tetrahymena thermophila ribozyme. The most prominent structural motif of the P4-P6 domain is the tetraloop-tetraloop receptor interaction which "clamps" the distal parts of its hairpin-like structure. Destabilization of the tertiary structure of the P4-P6 domain by perturbation of the tetraloop-tetraloop receptor interaction alters the Mg2+-mediated folding pathway. The folding hierarchy of P5c approximately P4-P6 > periphery > catalytic core that is a striking attribute of the folding of the wild-type RNA is abolished. The initial steps in folding of the mutant RNA are > or =50-fold faster than those of the wild-type ribozyme with the earliest observed tertiary contacts forming around regions known to specifically bind Mg2+. The interaction between the mutant tetraloop and the tetraloop receptor appears coincidently with slowly forming catalytic core tertiary contacts. Thus, the stability conferred upon the P4-P6 domain by the tetraloop-tetraloop receptor interaction dictates the preferred folding pathway by stabilizing an early intermediate. A sub-denaturing concentration of urea diminishes the early barrier to folding the wild-type ribozyme along with complex effects on the subsequent steps of folding the wild-type and mutant RNA.
Shcherbakova
Inna
I
Department of Biochemistry, Albert Einstein College of Medicine, Bronx, NY 10461, USA.
Brenowitz
Michael
M
D000818:Animals; D020029:Base Pairing; D001483:Base Sequence; D007700:Kinetics; D008274:Magnesium / Q000737:chemistry / Q000378:metabolism*; D008969:Molecular Sequence Data; D009154:Mutation; D009690:Nucleic Acid Conformation; D016337:RNA, Catalytic / Q000737:chemistry*; D016053:RNA, Protozoan / Q000737:chemistry*; D013050:Spectrometry, Fluorescence; D016808:Tetrahymena thermophila / Q000235:genetics*; D013816:Thermodynamics
D016337:RNA, Catalytic; D016053:RNA, Protozoan; D008274:Magnesium