Investigation of the dynamics and conformational changes of the U1A protein

Anunciado, Divina B.

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

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

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

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

Copyright and License Information

Copyright 2010 Divina B. Anunciado

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