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Structural and Dynamic Basis for Assembly of the HIV-1 TAR-Tat Ribonucleoprotein Complex

dc.contributor.authorDethoff, Elizabeth A.en_US
dc.date.accessioned2011-09-15T17:16:56Z
dc.date.availableNO_RESTRICTIONen_US
dc.date.available2011-09-15T17:16:56Z
dc.date.issued2011en_US
dc.date.submitteden_US
dc.identifier.urihttps://hdl.handle.net/2027.42/86484
dc.description.abstractThe transactivation response element (TAR), located at the 5' end of the HIV-1 genome, regulates the transcription elongation step of viral RNA. The TAR stem-loop binds the HIV viral transactivator protein (Tat), which enhances viral transcription by recruiting the positive transcription elongation factor b (P-TEFb). The TAR-Tat interaction and formation of the TAR/Tat/P-TEFb ribonucleoprotein (RNP) complex remains poorly understood from a structural and dynamic standpoint. To better understand the formation of this complex, we have studied the structural and dynamic features of free TAR to elucidate motions in both the bulge and apical loop that may be important for adaptive recognition of protein targets and, ultimately, for anti-viral therapeutic design. We used a combination of nuclear magnetic resonance (NMR), molecular dynamics (MD), and mutagenesis to characterize the structural dynamics of free TAR. Residual dipolar couplings (RDCs), motionally-decoupled 13C relaxation (R1, R2), and 13C R1ρ relaxation dispersion were used in combination to characterize global and local motions occurring on fast (ps to ns) and slow (us to ms) timescales. These results were compared to an MD simulation of TAR. Our results show that the apical loop and bulge act as independent dynamical centers, with the apical loop undergoing complex dynamics on the microsecond to picosecond timescale that may be important for adaptive recognition. We have characterized a lowly-populated structure of the loop that is otherwise “invisible” to traditional NMR techniques, representing the first-ever characterization of an “invisible” state of an RNA loop by NMR. Overall, our results establish a broadly useful approach for characterizing the dynamics of RNA loops and shed light on the relationship between dynamics and biological function for the TAR element in the HIV genome.en_US
dc.language.isoen_USen_US
dc.subjectNMRen_US
dc.subjectRNAen_US
dc.subjectRDCen_US
dc.subjectRelaxationen_US
dc.subjectHIV-1en_US
dc.titleStructural and Dynamic Basis for Assembly of the HIV-1 TAR-Tat Ribonucleoprotein Complexen_US
dc.typeThesisen_US
dc.description.thesisdegreenamePhDen_US
dc.description.thesisdegreedisciplineChemistryen_US
dc.description.thesisdegreegrantorUniversity of Michigan, Horace H. Rackham School of Graduate Studiesen_US
dc.contributor.committeememberAl-Hashimi, Hashim M.en_US
dc.contributor.committeememberBrooks Iii, Charles L.en_US
dc.contributor.committeememberEngelke, David R.en_US
dc.contributor.committeememberRamamoorthy, Ayyalusamyen_US
dc.subject.hlbsecondlevelBiological Chemistryen_US
dc.subject.hlbsecondlevelChemistryen_US
dc.subject.hlbtoplevelScienceen_US
dc.description.bitstreamurlhttp://deepblue.lib.umich.edu/bitstream/2027.42/86484/1/dethoffe_1.pdf
dc.owningcollnameDissertations and Theses (Ph.D. and Master's)


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