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Dissertation/Thesis Abstract

Protein Function Study by NMR Spectroscopy
by Amero, Carlos Daniel, Ph.D., The Ohio State University, 2008, 244; 10630815
Abstract (Summary)

It is essential to understand how proteins perform their work and how they interact with other molecules. We have used Nuclear Magnetic Resonance (NMR) spectroscopy to study structure, dynamics and interactions in several systems: Poliovirus 3C protein, Cre recombinase, an archaeal RNase P protein (Rpp21) and Peptide Deformylase (PDF). This dissertation discusses these different biological systems in separated chapters. Chapter 1 serves as an introduction to the basics of Protein NMR.

Chapter 2 involves analysis of RNA interactions with the poliovirus 3C protein. Replication of picornaviral genomes requires recognition and binding by the virus-encoded 3C protein of at least three RNA elements. We have used chemical shift mapping to provide unique site-specific insight into residues of 3C that interact with RNA, and set the stage for detailed structural investigation of the 3C-RNA complex by NMR.

Chapter 3 is devoted to Cre recombinase from bacteriophage P. This enzyme recognizes specific DNA sequences on two DNA strands and mediates recombination between these sites via the formation of a covalent protein-DNA complex and a Holliday junction intermediate. We have used heteronuclear NMR methods to obtain insights into the free structure, dynamics and interaction with DNA.

Chapter 4 focuses in RNase P, the ubiquitous ribonucleoprotein enzyme responsible for cleaving the 5'-leader sequence of precursor tRNA (pre-tRNA) molecules during maturation. We have determined the solution structure of the Rpp21 protein from the hyperthermophilic archaeon, Pyrococcus furiosus (Pfu) using conventional and paramagnetic NMR techniques and characterize the interaction of Rpp21 with Pfu Rpp29.

Chapter 5. Peptide deformylase (PDF) is an enzyme that is responsible for removing the formyl group from nascently synthesized polypeptides in bacteria. It has attracted much recent attention as a potential target for novel antibacterial agents. We used 15N NMR spectroscopy and isothermal titration calorimetry to investigate the high-affinity interaction of PDF with actinonin, a naturally occurring potent PDF inhibitor. The results of these studies improve our understanding of the thermodynamic global minimum and has potentially important implications for structure-based design of protein-binding ligands.

Indexing (document details)
Advisor: Foster, Mark
Commitee: Foster, Mark, Grandinetti, Philip, Dr, Wu, Justin, Dr
School: The Ohio State University
Department: Biophysics
School Location: United States -- Ohio
Source: DAI-B 78/11(E), Dissertation Abstracts International
Subjects: Molecular biology, Biochemistry, Biophysics
Keywords: Cre recombinase, Dynamics, Interaction with DNA, NMR, Poliovirus 3C protein, Structure
Publication Number: 10630815
ISBN: 978-0-355-01156-2
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