Date of Award

2016

Document Type

Open Access Dissertation

Department

Chemistry and Biochemistry

Sub-Department

College of Arts and Sciences

First Advisor

Qian Wang

Abstract

The complexity of cellular environments presents a challenge to efforts at studying and elucidating biomolecules in cells. Covalent ligation of biomolecules with fluorescent molecules that have a reactive functional group that can react with a complementary functional group on biomolecules is one way that has been utilized to study biomolecules. It enables visualization and tracking of ligation and transportation of molecules within the cell. However, the myriad of functional groups within the cell makes achieving selectivity difficult. To circumvent this problem, reactions that do not interfere with biology have been developed. These reactions, referred to as bioorthogonal reactions must be rapid, selective to the reactive species in the cellular environment and non-toxic to the cells. In Chapter 1 of this work, we discussed some of the key reactions developed to date and their applications in bioconjugation.

In Chapter 2, we investigated the regioselectivity of secondary amine-catalyzed inverse electron demand Diels-Alder reaction (iDA) of unsymmetrical tetrazines with aldehyde or ketones. Only one regioisomer was observed for the reactions of the unsymmetrical tetrazines with aldehydes and ketones. In addition, fluorogenic dyes were developed with tetrazine as the “triggering group”. The reaction was amenable to aqueous environment and hence was used to label aldehydes in cells.

In Chapter 3, a quartz crystal microbalance (QCM)-based sensor for detecting aldehydes was developed. By immobilizing graphene followed by 1,8-diaminonapthalene (DAN) on a QCM chip, a sensitive and facile aldehyde and ketone sensor was developed. The binding of the aldehydes to the DAN on the QCM chip resulted in a decrease in the fundamental resonant frequency of the functionalized QCM chips. The magnitude of the frequency change is directly proportional to the mass added. The utility of the probe for labeling biomolecules was demonstrated by using as a sensor for sialic acids pretreated with sodium periodate at mild conditions. In addition, the sensor was used to detect sialic acid on a sialoprotein and on bone marrow mesenchymal stem cells.

In Chapter 4, pyrene-anchored boronic acids were designed, synthesized and subsequently used to label glycans on cells. The boronic acids bound to the diols of glycans whiles pyrenes allowed for fluorescence imaging and tracking of the molecule in the cell. We observed the binding of the boronic acids to the glycans allowed for transport of the pyrene-anchored boronic acids into the cytoplasm of the cell.

Available for download on Tuesday, December 12, 2017

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