Characterizing complex dynamics in the transactivation response element apical loop and motional correlations with the bulge by NMR, molecular dynamics, and mutagenesis.
The HIV-1 transactivation response element (TAR) RNA binds a variety of proteins and is a target for developing anti-HIV therapies. TAR has two primary binding sites: a UCU bulge and a CUGGGA apical loop. We used NMR residual dipolar couplings, carbon spin relaxation (R(1) and R(2)), and relaxation dispersion (R(1rho)) in conjunction with molecular dynamics and mutagenesis to characterize the dynamics of the TAR apical loop and investigate previously proposed long-range interactions with the distant bulge. Replacement of the wild-type apical loop with a UUCG loop did not significantly affect the structural dynamics at the bulge, indicating that the apical loop and the bulge act largely as independent dynamical recognition centers. The apical loop undergoes complex dynamics at multiple timescales that are likely important for adaptive recognition: U31 and G33 undergo limited motions, G32 is highly flexible at picosecond-nanosecond timescales, and G34 and C30 form a dynamic Watson-Crick basepair in which G34 and A35 undergo a slow (approximately 30 mus) likely concerted looping in and out motion, with A35 also undergoing large amplitude motions at picosecond-nanosecond timescales. Our study highlights the power of combining NMR, molecular dynamics, and mutagenesis in characterizing RNA dynamics.
Dethoff
Elizabeth A
EA
Department of Chemistry and Biophysics, University of Michigan, Ann Arbor, Michigan 48109-1055, USA.
Hansen
Alexandar L
AL
Musselman
Catherine
C
Watt
Eric D
ED
Andricioaei
Ioan
I
Al-Hashimi
Hashim M
HM
D001483:Base Sequence; D002247:Carbon Isotopes; D015497:HIV-1 / Q000737:chemistry*; D009682:Magnetic Resonance Spectroscopy; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016296:Mutagenesis; D009154:Mutation / Q000235:genetics; D009587:Nitrogen Isotopes; D009690:Nucleic Acid Conformation; D012367:RNA, Viral / Q000737:chemistry* / Q000235:genetics; D020218:Response Elements / Q000235:genetics*; D015533:Transcriptional Activation / Q000235:genetics*
D002247:Carbon Isotopes; D009587:Nitrogen Isotopes; D012367:RNA, Viral