| Following the discovery of deoxyribonucleic acid (DNA) and its role as the genetic carrier of life, researchers have investigated the potential use of DNA as a molecular tool in chemical biology. Along this line, my Ph.D. thesis examines the use of DNA oligonucleotides and peptide-oligonucleotide conjugates (POCs) as material to capture and organize biological macromolecules in nanotechnology and molecular biology.;Using POC molecules, we developed a new method of positioning proteins at addressable locations on DNA nanostructures. Described in chapter 2 is the design of a DNA nanostructure that displayed single-stranded DNA probes that were complimentary in sequence to an oligonucleotide conjugated to the c-myc peptide. When the DNA array was assembled, the c-myc POCs were organized on the array surface and remained accessible for recognition by the c-myc antibody. This research was the first demonstration of peptide nanoarrays using POCs to specifically organize proteins on DNA nanostructures.;In chapter 4, we created bivalent reagents (termed synbodies) using POC molecules composed of complimentary strands of DNA independently conjugated to two peptides that exhibited non-overlapping binding to a target protein. When the POCs are hybridized, bivalent reagents are created that display two peptides on a double-stranded DNA scaffold with varying spatial separation between the peptides. We have observed that the synbodies with the optimal peptide separation distance exhibit ∼1000-fold enhancement in binding affinity to the target protein in comparison to the peptides that comprise the bivalent reagents.;Alternatively, chapter 3 describes the use of in vitro evolution to select for single strand DNA molecules (termed aptamers) with discrete tertiary structures that are capable of specifically binding the histone H4 protein with a acetyl modification. When compared to a commercial antibody, the DNA aptamer binds with similar affinity, but much stronger specificity when probed with different modified and unmodified protein targets. This research demonstrated the ability of DNA aptamers to specifically recognize small chemical modifications among protein targets.;Together, these projects demonstrate the utility of DNA oligonucleotides and POC molecules as chemical reagents to precisely organize biological materials on the nanoscale and produce high affinity protein binding agents. |
