Research Topics
Department of Physics
Ph.D. (1996) University of California, Berkley
Research Topics
Single molecule biophysical tools applied to:
- Enzyme mechanism
- DNA - protein interaction
- Protein folding and RNA folding
- Membrane protein interactions
- Development of ultra-high resolution fluorescence microscopy
Research Interests
A complete understanding of the bio-molecular functions demands an understanding of the internal motion of the molecules in relation to its function. New single-molecule methodologies allow us to look at the real-time structural changes during biological events, probing the heart of the structure-function relationship. The focus of my research is the application of single molecule fluorescence microscopy and single-molecule manipulation tools to directly probe biological processes. Another area of interest is the marriage of two new powerful techniques: single-molecule methods and DNA array-based methods.
1) Conformational Dynamics of Novel RNA Structures RNA plays important cellular roles in information storage, transfer and processing, in addition to serving as a catalytic core for multi-protein complexes such as ribosome. We have applied our single-molecule approaches to study how an RNA molecule folds into 3D structure and dynamically changes its shape spontaneously or in response to other biomolecules or ions. Future efforts will be focused on the dynamic structural changes of novel RNA motifs that are found in nature as part of larger ribonucleo-protein complexes such as ribosome and spliceosome.
2) Single-molecule Study of DNA Helicases Nucleic acid unwinding is an essential step for many bio-molecular processes. For instance, DNA has a double helix structure formed by two strands that must be 'unwound' before being copied. We successfully developed a unique single-molecule approach that can reveal the molecular mechanism of helicases that are not accessible by other conventional methods. We will apply novel single-molecule assays to investigate how helicases consume free energy released by breaking chemical bonds in the 'fuel' molecule and couple it to their structural changes and move along DNA molecule and unwind it. Innovative combinations of fluorescence and manipulation will be developed to study how the linear and torsional tension on the DNA influence the function of helicases.
3) Single-molecule Studies of Function, Structural Change and Interaction of Membrane Proteins Many important biological processes occur on membranes. Single-molecule fluorescence techniques are ideal for observing complex, multi-step processes of membrane proteins. Supported bilayers with reconstituted membrane protein, provides a clean model system. We are studying the various pathways of bilayer formation by vesicle fusion using single-vesicle assays. New forms of bilayers, combined with ultra-smooth glass support and polymeric interfaces will be developed to preserve the activity of trans-membrane proteins. The resulting membrane quality will be quantified by single-lipid diffusion analysis and fluorescence-based functional assays of membrane proteins. Then, we will proceed to study how membrane proteins are partitioned and fold in the membrane and how they are recruited and organized into a functional form upon external stimuli. Particular systems of interests are immunological synapse formation by T-cell and target cell, vesicle fusion induced by SNARE complexes, and membrane protein localization directed by translocases.
4) High-throughput Screening of Nucleic Acid Arrays Based on Single-molecule Observables Single-molecule assays and DNA array-based techniques are two new and powerful tools. We propose to combine the two and build arrays of unique nucleic acids structures that will be characterized by single-molecule assays. For instance, many nucleic acid structures can be built from two or more oligonucleotides that can be tethered to a surface in a step-wise manner via micro-fluidic delivery. Motorized stage movement synchronized with fluorescence data acquisition/analysis will allow us to rapidly characterize individual mutants based on their properties that can be classified only at the single-molecule level (for example, folding pathways, rare and transient reactions).
Key Words Single molecule, fluorescence
spectroscopy/microscopy, optical tweezers, magnetic tweezers, helicase, DNA
repair, DNA replication, RNA folding, membrane proteins, supported bilayers,
nanoparticles
Current Research Funding Start up and research board (UIUC)