Date of Award

Spring 2017

Document Type

Dissertation

Degree Name

Doctor of Philosophy in Chemistry - (Ph.D.)

Department

Biomedical Engineering

First Advisor

Edgardo Tabion Farinas

Second Advisor

Haidong Huang

Third Advisor

Xiaoyang Xu

Fourth Advisor

Tamara M. Gund

Fifth Advisor

Yong Ick Kim

Sixth Advisor

Pradyot Patnaik

Abstract

The biological effects of DNA alkyl adducts are difficult to evaluate at the cellular level due to their instability. Synthesis of oligonucleotides that contain a single N7-alkylguanine has become a vital tool to achieve the above goal. However, the instability of N7-alkyguanines is not compatible with the phosphoramidite chemistry used by solid-phase oligonucleotide synthesis either. Development of chemically stable analogues of unstable DNA lesions enables accurate study of polymerase bypass. The design and successful synthesis of N7-hydroxyethyl-9-deaza-2′-deoxyguanosine and N7-oxoethyl-9-deaza-2′-deoxyguanosine as the stable analogues of N7-hydroxyethyl- 2′-deoxyguanosine and N7-oxoethyl-2′-deoxyguanosine, respectively, are reported. The synthesis of these two nucleosides whose N7 side chains are protected by TBS for the convenience of conversion to phosphoramidites are also developed. The C-glycosidic bonds of these compounds are demonstrated to be stable under strong acidic and basic conditions. These analogues will become versatile tools to study the replication and repair of DNA alkylation damages.

DNA oxidation product 8-oxoGua has been suggested as a biomarker for early cancer diagnosis. An artificial receptor for the free base of 8-oxoGua on a triplex DNA backbone was previously developed. However, accurate detection of 8-oxoGua in urine samples was affected by the presence of a large excess of guanine. Herein, a unique strategy to convert such a receptor to a colorimetric biosensor by conjugating DNA strands to gold nanoparticles (GNP) is developed. Binding of 8-oxoGua to the receptor caused the conjugation of GNP, resulting in diagnostic red-to-purple color changes. The presence of multiple binding cavities enhances the binding-induced stabilization effect and widened the temperature window used for detection. By simply incubating our sensor with a sample, 8-oxoGua can be detected at submicromolar concentrations with a UV-vis spectrometer or even by naked eye. The detection limit in a urine matrix is determined as 126 nM and the response range covers a major portion of the biologically relevant concentration range.

Included in

Chemistry Commons

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