Research

Our laboratory is interested in developing tailor-made catalysts and sensors for biotechnology and molecular medicine. A key component of our research is the use of laboratory evolution strategies to evolve proteins and nucleic acids with novel structures and functions. We apply a chemical biology approach that combines traditional synthetic organic chemistry and molecular biology with genetics, proteomics, materials science, and structural biology to solve problems of fundamental and practical importance. Specific projects in our laboratory include:

Evolution of Novel Protein Folds and Functions: Whether nature uses all or most of the possible protein folds or just a subset of all possible protein folds remains an unanswered question in biology. Current databases of biological protein structures estimate the total number of unique single-domain protein folds to be at or near one thousand. Given the vast size of the protein universe, it seems likely that examples should exist where protein sequences from non-biological sources could fold to adopt three-dimensional structures that are physically realistic, but topologically different than proteins found in nature. To address this problem, we apply a cell-free technology called mRNA display to select and optimize functional proteins from large libraries (≥10 trillion) of random amino acid sequences. The goal of this study is to identify polypeptides capable of adopting discretely folded structures with pre-defined functions. By solving the complete three-dimensional structure of these in vitro evolved non-biological proteins, we can begin to examine the extent to which nature samples the protein universe. Shown below are a few examples of the proteins that we have discovered by this process and protein crystals used to obtain X-ray diffraction data.



Design of Artificial Affinity Reagents for Proteomic Research: The ability to monitor disease-causing agents (biomarkers) in human saliva or blood holds great promise for the detection and treatment of many diseases, including cancer. Although most analytical techniques for detecting protein levels in human samples currently rely on ELISA-based assays or mass spectroscopic analysis, protein capture arrays have gained considerable interest as a future technology for proteome-wide analysis. In collaboration with Profs. Neal Woodbury and Stephen Johnston, we are beginning to develop a protein detection system that relies on synthetic antibodies (synbodies) to detect and profile disease-related proteins. Our approach to this problem involves a high throughput screen for ligand discovery followed by a combinatorial linking strategy for synbody assembly. Artificial antibodies produced in this manner have been shown to function with high affinity and specificity in solution and on a solid surface, and have been used successfully as protein affinity reagents in a recognition imaging experiment on an atomic force microscope (AFM).

Nanometer-Scale Inorganic Assemblies: In the race to create computer chips with smaller and smaller feature sizes, new bottom-up strategies are beginning to emerge that might one day surpass traditional chip making techniques. In collaboration with Prof. Hao Yan, we have developed a general approach for producing inorganic arrays with nanometer-scale features. Our approach relies on a new technology, called nanodisplay, which positions peptides known to bind and/or nucleate inorganic nanocrystals at specific sites on a DNA nanoarray. To expand the number of peptide-inorganic pairs used in this self-assembly process, we are currently evolving new peptides that bind inorganic materials with metallic, magnetic, acoustic, conducting, and semi-conducting properties. Shown here is one of our in vitro selected peptides that has been fluorescently labeled and bound to a ZnS nanocrystal. The contrasting green and black regions highlight the specificity of the selected peptide for individual faces of the inorganic lattice.

Selection of Unnatural Nucleic Acid Sensors and Catalysts: Threose nucleic acid (TNA) is a four-carbon sugar analogue of RNA that exhibits complementary Watson-Crick base pairing with DNA, RNA, and other TNA oligonucleotides. This unusual property together with the chemical simplicity of threose, suggest that TNA might be a progenitor candidate of RNA. To examine this hypothesis in greater detail, we are developing an in vitro selection strategy for evolving TNA aptamers and enzymes. By comparing the functional properties of TNA with RNA, we aim to determine the fitness of TNA as an alternative genetic material. Apart from our interest in generating alternative genetic systems, we are also exploring the possibility of using TNA as a sensor for in vivo imaging applications.