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    Design and evolution of functional nucleic acids

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    levym032.pdf (5.456Mb)
    Date
    2003
    Author
    Levy, Matthew
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    Abstract
    Functional nucleic acids provide insight into the ‘RNA world,’ the proposed period in Earth’s history where RNA served both as catalyst and as the genetic material. Additionally, they are potentially valuable tools for biotechnical applications. Like catalytic RNA, DNA has proven capable of catalyzing chemical reactions. Using a novel ligation chemistry, we have employed in vitro selection to isolate a catalytic DNA molecule (deoxyribozyme) that is capable of forming an unnatural nucleotide linkage. In addition, we have used a combination of rational design and in vitro selection to construct effector-dependent deoxyribozymes that are allosterically activated by the small molecule ATP. These results further strengthen the idea that life arose from a nucleic acid based metabolism and also bode well for the design of more stable nucleic acid based analyte detection systems. In addition to the design of novel DNA catalysts, we have been interested in the origin of the first simple self replicating systems. Simple replicators based on short oligonucleotides as well as peptides have been demonstrated. We have designed a cross-catalytic system based on a well characterized peptide-RNA aptamer interaction, in which the peptide serves as a template for the ligation of RNA aptamer half-molecules. Our results demonstrate that the peptide could specifically enhance the rate of RNA ligation, and suggest the possibility for increased diversity of early replicators. We have also designed an autocatalytic system based on the fast and efficient RNA cleaving 10-23 deoxyribozyme. In this system, two complementary deoxyribozymes have been inactivated by circularization. The circular deoxyribozymes are capable of serving as substrates for the linear enzymes such that linearization results in a cascade of cleaving reactions that exhibits exponential growth. A selection scheme based on this reaction resulted in optimal sequence selection and demonstrates the first in vitro selection experiment conducted in the absence of proteins.
    Department
    Institute for Cellular and Molecular Biology
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    text
    URI
    http://hdl.handle.net/2152/1106
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    © The University of Texas at Austin