INVESTIGATION OF BIOMOLECULAR INTERACTIONS FOR DEVELOPMENT OF SENSORS AND DIAGNOSTICS

• Main Author: Zhang, Xiaojuan
• Format: Others
• Published: VCU Scholars Compass 2011
• Subjects: AFM; Biomolecular interaction; Biosensing; Aptamer; Engineering
• Online Access: http://scholarscompass.vcu.edu/etd/294; http://scholarscompass.vcu.edu/cgi/viewcontent.cgi?article=1293&context=etd

The highly specific recognition processes between biomolecules mediate various crucial biological processes. Uncovering the molecular basis of these interactions is of great fundamental and applied importance. This research work focuses on understanding the interactions of several biomolecular recognition systems and processes that can provide fundamental information to aid in the rational design of sensing and molecular recognition tools. Initially, a reliable and versatile platform was developed to investigate biomolecular interactions at a molecular level. This involved several techniques, including biomolecule functionalization to enable attachment to self-assembled monolayers as well as atomic force microscopy (AFM) based force spectroscopy to uncover the binding or rupture forces between the receptor and ligand pairs. It was shown that this platform allowed determination of molecular binding between single molecules with a high specificity. The platform was further adapted to a general sensing formulation utilizing a group of flexible and adaptive nucleic acid recognition elements (RNA and DNA aptamers) to detect specific target proteins. Investigation of interactions at the molecular level allowed characterization of the dynamics, specificity and the conformational properties of these functional nucleic acids in a manner inaccessible via traditional interaction studies. These interactions were then adapted to aptamer-based detecting methods that at the ensemble or bulk scale, specifically taking advantage of mechanisms uncovered in the biophysical study of this system. A quartz crystal microbalance (QCM) was used to detect protein targets at the bulk level and the affinities and binding kinetics of these systems were analyzed. Along with AFM-based force spectroscopy, ensemble-averaging properties and molecular properties of these interactions could be correlated to contribute to bridging the gap across length scales. Finally, more broadly applicable sensing platform was developed to take advantage of the unique properties of aptamers. DNA was employed both as a carrier and as a molecular recognition agent. DNA was used as a template for nanoconstruction and fabricating unique shapes that could enhance the aptamer-based molecular recognition strategies. With aptamers tagged to distinct nanoconstructed DNA, a novel shape-based detecting method was enabled at the molecular level. The results demonstrated that this is a flexible strategy, which can be further developed as ultrasensitive single molecule sensing strategy in complex environments.