Title: MEMS-based temperature-dependent characterization of biomolecular interactions
Description: Biomolecular interactions are of fundamental importance for a wide variety of biological processes. Temperature dependence is a ubiquitous effect for biomolecular interactions as most biological processes are thermally active. Understanding the temperature dependence of biomolecular interactions is hence critical for a wide variety of applications in fundamental sciences and drug discovery and biotherapeutics. Micro-Electro-Mechanical Systems (MEMS) technology holds great potential in facilitating temperature-dependent characterization of biomolecular interactions by providing on-chip microfluidic handling with drastically reduced sample consumption, and well-controlled micro- or nanoscale environments in which biomolecules are effectively manipulated and analyzed. This thesis is focused on various MEMS-based devices for temperature-dependent characterization of biomolecular interactions. Biomolecular interactions can occur with biomolecules in solution or with either the target or receptor molecules immobilized to a solid surface.

For surface-based biomolecular interactions, we first present microcantilever-based characterization of biomolecular affinity binding with in-situ temperature sensing, using a demonstrative system of platelet-derived growth factor (PDGF) and an inhibitory ligand. The temperature-dependent kinetic and equilibrium binding properties are determined. In addition, a microfluidic approach for temperature-dependent biomolecular behavior with single-molecule resolution is also presented. Using a platform that combines microfludic sample handling, on-chip temperature control, and total internal reflection fluorescence (TIRF) microscopy, we have studied the temperature dependence of the structural dynamics of transfer RNA (tRNA) translocation through ribosome in protein synthesis. For solution-based biomolecular interactions, we mainly focus on calorimetry, a technology that directly measures heat evolved in biological processes. We first present a MEMS differential scanning calorimetric (DSC) sensor integrating highly sensitive thermoelectric sensing and microfluidic handling for thermodynamic characterization of biomolecules.

We have characterized the unfolding of protein (e.g. lysozyme) at minimized sample consumption with thermodynamic properties determined, including the specific heat capacity, molar enthalpy change, and melting temperature. In addition, we also present the development of a variant of standard DSC, temperature-modulated DSC (AC-DSC), on a MEMS device for thermodynamic characterization of biomolecules. Preliminary results again with lysozyme unfolding at optimum modulation frequencies have been presented with thermodynamic properties determined. Furthermore, we have developed a MEMS isothermal titration calorimeter (ITC) integrating thermally isolated calorimetric chambers, on-chip passive mixing, and environmental temperature control, for temperature-dependent characterization of biomolecular interactions. We have characterized the interactions of 18-Crown-6 and barium chloride, as well as ribonuclease A and cytidine 2'-monophosphate, in a 1-µL volume with low concentrations (ca. 2 mM). Thermodynamic properties, including the stoichiometry, equilibrium binding constant, and enthalpy change, are also determined.