The School of Molecular and Cellular Biology at the University of Illinois at Urbana-Champaign

We are interested in elucidating the role of metal ions in proteins and catalytic DNA/RNA (DNA and RNA with catalytic functions), designing metalloenzymes with novel structures and functions, and exploring the use of the enzymes in biotechnological and pharmaceutical applications. To achieve these goals, we are developing new approaches that combine the advantages of both chemical and biochemical systems.

Metalloprotein Design and Engineering. Metalloproteins are an important member of protein family that can catalyze some of the most difficult reactions with fine control of reactivity and selectivity. While much progress has been made on the study of native metalloproteins, little is known about how to design a metalloprotein with desired structure and function. We have been using stable, easy-to-produce, and well-characterized proteins as a scaffold for designing and engineering artificial metalloproteins that either have similar structural and functional properties of much more complex native proteins, or possess new structure or reactivity that is unprecedented in nature. We have designed and engineered a copper center into azurin and myoglobin that mimics the CuA and CuB site, respectively, in cytochrome oxidase (a terminal oxidase of the aerobic respiratory chain). We have also designed and engineered a manganese-binding site in cytochrome c peroxidase that closely resembles the manganese-binding site in manganese peroxidase (a heme enzyme that has great promise in providing renewable energy and in destroying environmental pollutants). Through rational design, cytochrome c peroxidase was also redesigned to closely mimic a cytochrome P450, a heme enzyme that is responsible for many regio-, stereo- and enantio-selective chemical transformations in biological system and in chemical synthesis.

New DNA and RNA Enzymes as Anti-viral Pharmaceutical Agents or as Metal Biosensors. Recent results have shown that DNA and RNA not only are important in genetic information transfer, but also can be enzymes that can catalyze a variety of different reactions. This discovery has lead to the development of catalytic DNA/RNA as promising pharmaceutical agents against AIDS and other retroviral diseases. To obtain new DNA and RNA enzymes with high anti-viral activity for pharmaceutical applications and with strong metal-binding affinity for biophysical studies, we have used a technique called in vitro selection to select, from a library of 1014-1015 DNA molecules with different sequences, new catalytic DNAs with high efficiency and strong metal ion affinity. These results open a new avenue for spectroscopic and kinetic study of catalytic metal binding sites in nucleic acid enzymes and for their pharmaceutical applications. Furthermore, we are the first to report a new application of catalytic DNA as biosensors for metal ions such as Pb2+. The DNA biosensor combines the high selectivity of catalytic DNA with the high sensitivity of fluorescence detection, and can be applied to the quantitative detection of metal ions over a wide concentration range. The easy adaptability of DNA to optical fiber and chip technology makes this DNA enzyme system an ideal choice for simple, real-time, and remote sensing of metal ions in applications such as environmental monitoring, clinical toxicology, and industrial process monitoring.