University of Illinois Urbana-Champaign

Investigation of the dynamics and conformational changes of the U1A protein

Anunciado, Divina B.

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Anunciado_Divina.pdf

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https://hdl.handle.net/2142/18490

Description

Title
Investigation of the dynamics and conformational changes of the U1A protein
Author(s)
Anunciado, Divina B.
Issue Date
2011-01-21T22:42:38Z
Director of Research (if dissertation) or Advisor (if thesis)
Baranger, Anne M.
Doctoral Committee Chair(s)
Baranger, Anne M.
Committee Member(s)
  • Gruebele, Martin
  • van der Donk, Wilfred A.
  • Zhao, Huimin
Department of Study
Chemistry
Discipline
Chemistry
Degree Granting Institution
University of Illinois at Urbana-Champaign
Degree Name
Ph.D.
Degree Level
Dissertation
Date of Ingest
2011-01-21T22:42:38Z
Keyword(s)
  • conformational changes
  • dynamics
  • RNA-recognition motif
  • cooperative interactions
  • time-resolved fluorescence anisotropy
  • temperature-jump kinetics
  • stopped-flow kinetics
Abstract
The U1A-SL2 RNA complex is a model system for studying interactions between RNA and the RNA recognition motif (RRM), which is one of the most common RNA binding domains. Despite extensive studies conducted to study the origins of U1A-RNA affinity and specificity, little is known about the kinetics and mechanism of binding. This dissertation aims to help understand the contributions of dynamic processes to the U1A-RNA complex formation. The first chapter introduces the significance of studying RNA-binding proteins, and RNA-Recognition Motif proteins in particular. It also presents the general dynamic processes that have been observed in RRMs. Chapter 2 is about the characterization of the dynamics of an essential helix in the U1A protein, helix C. The time-resolved anisotropy data obtained here supports the induced fit mechanism of binding. Chapter 3 describes the kinetic studies of dissociation of the U1A-SL2 RNA complex using laser T-jump and stopped-flow fluorescence methods. Analysis of the kinetic data suggested three phases of dissociation with two intermediate states. Chapter 4 is a follow up study of chapters 2 and 3 to further investigate helix C dynamics. The experimental design makes use of tryptophan fluorescence quenching with histidine and cysteine. Chapter 5 explores the existence of cooperative interactions in the U1A protein. Taken together, all these results begin to build a comprehensive picture of the complex dynamic processes involved in the formation of an RRM-RNA complex.
Graduation Semester
2010-12
Permalink
http://hdl.handle.net/2142/18490
Copyright and License Information
Copyright 2010 Divina B. Anunciado

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