Dr. Thomas P. ShieldsOffice: Center for Science and Technology - 491Phone: (619) 260-7527 e-mail Address: tshields@sandiego.edu Personal website: www.sandiego.edu/~tshields |
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Dr. Thomas P. ShieldsOffice: Center for Science and Technology - 491Phone: (619) 260-7527 e-mail Address: tshields@sandiego.edu Personal website: www.sandiego.edu/~tshields |
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Academic Background
Bachelor of Science: University of Notre
Dame 1989. Major : Chemistry
Doctor of Philosophy: California Institute of Technology 1995; Advisor : Dr. Jacqueline K. Barton
Postdoctoral Research: University of Colorado at Boulder 1995-1998; NMR of RNA Aptamers; Advisor: Dr. Art Pardi
Courses Taught
Chemistry 10A General Chemistry
Chemistry 11A General Chemistry Laboratory
Chemistry 130 Biochemistry
RNA structure & function; In vitro selected RNAs, RNA - drug interactions.
Description of Research
In vitro selection is a powerful combinatorial technique capable of isolating RNA molecules with a particular binding affinity or catalytic activity from an initial randomized pool of RNA sequences. High affinity RNA sequences, termed aptamers, have been isolated for a diverse array of targets, including both proteins and small molecules.
Since RNA aptamers display high affinity and high selectivity for their target ligands, they are exciting candidates for development into clinical biosensors. Two well-studied RNA aptamers that bind small molecule targets are the RNAs that bind either theophylline (an asthmatic drug) or ATP (the ubiquitous "currency" of biochemistry). In both cases, addition of the target to the RNA aptamer leads to formation of a well-defined tertiary complex. These RNA aptamers serve as model systems in our lab to explore new biosensor designs.
Pseudouridine and 1-methyladenosine are two modified nucleosides that are present in cellular RNA. These molecules are markers for cellular turnover, and therefore are important prognosticators for cancer since their urinary levels increase in parallel with the disease state. Quantitation of these modified nucleosides in the milieu of other urinary cellular byproducts is a challenging task most often approached by HPLC analysis. An in vitro selected RNA molecule specific for pseudouridine or 1-methyladenosine could be the basis for a simple biosensor for both cancer prognosis and monitoring chemotherapy.
The research will progress along two parallel fronts. In one project, in vitro selection will identify RNA molecules capable of binding to pseudouridine and 1-methyladenosine with high affinity and selectivity from an initial pool of random RNA sequences. In parallel, two well-characterized RNA aptamers will be converted into simple biosensors for their targets. The new RNA aptamers for pseudouridine and 1-methyladenosine will then be converted into biosensors using the technologies developed for the well-charactersized RNA aptamers.
The basis for our biosensor is the hybridization of an RNA aptamer to a complementary fluorescent DNA. The formation of the RNA-DNA hybrid duplex causes a decrease in the fluorescence of the fluorescent DNA strand. In the presence of the target drug (D), the RNA aptamer will preferentially bind to the drug, releasing the fluorescent DNA probe and regaining fluorescence intensity.
Publications
8. Zimmermann, G.R., Wick, C.L., Shields, T.P., Jenison, R.D. and Pardi, A (2000), Molecular Interactions and Metal Binding in the Theophylline-Binding Core of an RNA Aptamer. RNA 6, 659-667.
7. Shields, T.P., Mollova, E., Ste. Marie, L.J., Hansen, M.R. and Pardi, A.. (1999), High-Performance Liquid Chromatography Purification of Homogenous-Length RNA Produced by trans Cleavage with a Hammerhead Ribozyme. RNA 5, 1259-1267.
6. Zimmermann, G.R., Shields, T.P., Jenison, R.D., Wick, C.L. and Pardi, A. (1997), A Semiconserved Residue Inhibits Complex Formation by Stabilizing Interactions in the Free State of a Theophylline-Binding RNA. Biochemistry 37, 9186-9192.
5. Shields, T.P. and Barton, J.K. (1995), Sequence-Selective DNA Recognition and Photo-cleavage: A Comparison of Enantiomers of [Rh(en)2phi]3+. Biochemistry 34, 15037-15048.
4. Shields, T.P. and Barton, J.K. (1995), Structural Examination of Enantioselective Intercalation: 1H NMR of [Rh(en)2phi]3+ Isomers Bound to d(GTGCAC)2. Biochemistry 34, 15049-15056.
3. Collins, J.G., Shields, T.P. and Barton, J.K. (1994) NMR Evidence for Classical Intercalation By a Non-Classical Octahedral Metal Complex. J. Am. Chem. Soc. 116, 9840.
2. Krotz, A.H., Kuo, L.Y., Shields, T.P. and Barton, J.K. (1993) DNA Recognition By Rhodium(III) Polyammine Intercalators : Considerations of Hydrogen Bonding and Van Der Waals Contacts. J. Am. Chem. Soc. 115, 3877.
1. Schaefer, W. P., Krotz, A. H., Kuo, L.Y., Shields, T. P. and Barton, J. K. (1992), Metallo-intercalators: Structure of rac -Bis(ethylenediammine)(9,10-phenanthrenequinone diimine) rhodium(III) Tribromide Trihydrate (rac -[Rh(en)2phi]Br3 . 3 H2O). Acta Cryst. C48, 2071.
