Emeritus Professor
Hewett University Professor
Emeritus Professor <br />Hewett University Professor Profile Image

Department of Chemistry
University of Nebraska-Lincoln
737 Hamilton Hall
Lincoln, NE 68588-0304
(402) 472-3316
lparkhurst1@unl.edu

Education

Ph.D., Yale University
M.S., Yale University
B.S., Yale University

Research Interests

Biomolecular sciences, laser chemistry

Current Research

Our lab is working to understand fundamental processes in the initiation of eukaryotic transcription. Our goal is to understand the structural and energetic features that control the recognition of DNA promoter sequences by TBP (TATA Binding Protein) and the further assembly of the multi-protein transcription initiation complex. We use numerous approaches that include microcalorimetry, steady-state and time-resolved fluorescence and anisotropy in the psec-nsec time regime, stopped-flow rapid reaction kinetics, and molecular dynamic studies of protein-DNA interactions using the university supercomputer.

Figure 1

Using these various techniques, we have shown that the binding of DNA by TBP is a multi-step process, depicted below, in which DNA is strongly bent (by about 80o) in the very first detectable complex, one that we have proposed is the crucial intermediate involved in building the pre-initiation complex. We have used time-resolved fluorescence energy transfer (FRET) to show that the bending of the DNA is sequence dependent and correlates with the activity of the promoter in transcription.

Figure 2

Using pulsed laser techniques, we have developed time-resolved acceptor FRET to measure distances in protein complexes in solution that exceed 100. At right is the ternary complex of TBP-DNA-TFIIA, with the distance between arrows at sites "A" and "B" corresponding to 115, a distance we have measured between dyes fluorescein (F) and xRhodamine (xR) attached to the ends of a 29-mer duplex DNA.

Figure 3

For more information, please visit the Parkhurst Research Group Homepage.

Selected Publications

(1) A small molecule directly inhibits the p53 transactivation domain from binding to replication protein A By: Glanzer, Jason G.; Carnes, Katie A.; Soto, Patricia; et al. NUCLEIC ACIDS RESEARCH Volume: 41 Issue: 3 Pages: 2047-2059 Published: FEB 2013

(2) TATA-Binding Protein Recognition and Bending of a Consensus Promoter Are Protein Species Dependent, Whittington, J. E., Delgadillo, R. F., Attebury, T., Parkhurst, L. K., Daugherty, M. A. and Parkhurst, L. J., Biochemistry (2008), published on the Web 06/14/2008.

(3) Changes in DNA bending and flexing due to tethered cations detected by fluorescence resonance energy transfer Williams, Sarah L.; Parkhurst, Laura K.; Parkhurst, Lawrence J.. Department of Chemistry, University of Nebraska-Lincoln, Lincoln, NE, USA. Nucleic Acids Research (2006), 34(3), 1028-1035.

(4) Native Human TATA-binding Protein Simultaneously Binds and Bends Promoter DNA without a Slow Isomerization Step or TFIIB Requirement K. M. Masters, K. M. Parkhurst, M. A. Daugherty, L. J. Parkhurst, Journal of Biological Chemistry 2003, 278, 31685-31690.

(5) Comparison of TATA-binding protein recognition of a variant and consensus DNA promoters R. M. Powell, K. M. Parkhurst, L. J. Parkhurst, Journal of Biological Chemistry 2002, 277, 7776-7784.

(6) Time-resolved fluorescence resonance energy transfer studies of DNA bending in double-stranded oligonucleotides and in DNA-protein complexes L. J. Parkhurst, K. M. Parkhurst, R. Powell, J. Wu, S. Williams, Biopolymers 2002, 67, 180-200.

(7) DNA bends in TATA-binding protein.TATA complexes in solution are DNA sequence-dependent J. Wu, K. M. Parkhurst, R. M. Powell, M. Brenowitz, L. J. Parkhurst, Journal of Biological Chemistry 2001, 276, 14614-14622.

(8) DNA sequence-dependent differences in TATA-binding protein-induced DNA bending in solution are highly sensitive to osmolytes J. Wu, K. M. Parkhurst, R. M. Powell, L. J. Parkhurst, Journal of Biological Chemistry 2001, 276, 14623-14627.

(9) Marked stepwise differences within a common kinetic mechanism characterize TATA-binding protein interactions with two consensus promoters R. M. Powell, K. M. Parkhurst, M. Brenowitz, L. J. Parkhurst, Journal of Biological Chemistry 2001, 276, 29782-29791.