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Sequence-specific minor groove binding polyamides : DNA recognition and applications

by Carter, Scott Reynolds

Abstract (Summary)
NOTE: Text or symbols not renderable in plain ASCII are indicated by [...]. Abstract is included in .pdf document. Pyrrole-imidazole polyamides bind to the minor groove of DNA with high affinity and specificity. This thesis explores the scope of this DNA recognition motif both in terms of improved recognition and application to problems of DNA detection and purification. Chapter 2 describes conjugates of polyamides with fluorescent dyes. Three cyanine dye butyric acids were synthesized and linked to the polyamide, [...]. These conjugates retain high specificities and high affinities for targeted DNA sequences, and show large fluorescence enhancements upon binding to double stranded DNA. Chapter 3 describes the use of polyamide conjugates as affinity reagents for the purification of double stranded DNA. Specifically, systems composed of biotin and histidine tagged conjugates that can rapidly and efficiently capture restriction fragment DNA are described. Chapter 4 focuses on the design and synthesis of constrained [...]-aminobutyric acid analogs designed to improve hairpin polyamide binding affinities through reduction of the entropic cost of binding to DNA. Six derivatives were synthesized as Boc protected monomers and were incorporated into polyamides by solid phase synthesis. The affinity and specificity of the compounds were analyzed by quantitative DNAse I footprinting. Chapter 5 describes preliminary synthetic efforts towards the recognition of G rich DNA sequences with polyamides containing imidazole dimers linked by alkyl chains rather than amides. Regioselective synthesis of an alkyl imidazole dimer suitably protected for solid phase polyamide synthesis is described. This dimer was incorporated into 2 polyamides for the recognition of 5'-WGGGGW-3' sequences. Alkyl substituted polyamides bound comparably to the corresponding amide linked compounds. An appendix summarizes work on the recognition of 5'-WGWGW-3' sequences by pyrrole-imidazole polyamides. [...] and [...] recognize a [...] binding site. Chapter 6 describes studies of the non-natural base pair between isoguanosine and isocytosine. A synthesis of deoxyisoguanosine (d-isoG) suitably protected for solid phase oligonucleotide synthesis is presented. Transcriptional studies on templates containing diosG showed that it directed the transcription of uridine rather than isoC, providing evidence that isoG can access different tautomeric forms.
Bibliographical Information:

Advisor:Peter Dervan

School:California Institute of Technology

School Location:USA - California

Source Type:Master's Thesis

Keywords:chemistry

ISBN:

Date of Publication:12/16/1998

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