This award is funded under the American Recovery and Reinvestment Act of 2009 (Public Law 111-5).
Gene-regulation by DNA-binding proteins is vital for living systems. Such proteins control gene expression by binding to particular DNA sequences, and the association must be rapid for quick response to cellular signals. The overall objective of this project is to obtain a physical understanding of the target search process whereby DNA-binding proteins rapidly and efficiently locate their target DNA sites in the presence of an enormous background of nonspecific DNA. In particular, the research component of this three-year project focuses on the structural, dynamic, and kinetic aspects of the target search by a gene-regulatory protein with multiple zinc-finger DNA-binding domains (DBDs). Using NMR spectroscopy, this project will analyze the target search by the Zif268 protein that contains three zinc-finger DBDs as a model system. This protein is an inducible gene-regulatory protein involved in neuronal plasticity and memory formation, and must efficiently find its target DNA sequences to mediate rapid cellular responses to neuronal signals. The Zif268 protein is an ideal model for the study of the target search process, because it has been extensively characterized in biochemical, structural and functional terms, and also exhibits excellent NMR spectra for both specific and non-specific DNA complexes. This project will address three major questions: 1) In terms of structure and dynamics, how does the Zif268 protein use the three DBDs for the target search while it moves on the nonspecific DNA 2) How efficient is the so-called direct transfer mechanism for the Zif268 protein 3) What is the relative contribution of the sliding mechanism to the target search at high DNA concentrations To answer these questions, various NMR approaches that have been developed in their previous studies will be used. This project will provide important and general insights into the target search by zinc-finger proteins, the most abundant class of eukaryotic gene-regulatory proteins. Experimental data obtained in this project will immediately be shared and discussed with Dr. Yaakov Levy's group (Weizmann Institute of Science, Israel), which has been studying theoretical and computational aspects of the facilitated target location by DNA-binding proteins. This international collaboration will integrate experimental and theoretical approaches to lead to more realistic views of the target search process.
Broader Impacts: By integrating research and education, this project aims to establish an institutional environment where students and researchers can effectively take advantage of NMR spectroscopy in their research. The education component offers opportunities whereby graduate students and research staff in the University of Texas Medical Branch (UTMB) can learn NMR as a research tool. The principal investigator (PI) of this project will design Mathematica-based syllabus for NMR spectroscopy, and also initiate an elective course to teach practical aspects of NMR spectroscopy, demonstrating actual data collection and analysis. Furthermore, the PI will continue to organize the NMR seminar as an effective means to share knowledge and tools. The educational component of this project includes science education for high school students and international student exchange with the collaborators laboratory. Through UTMB's HSSRP program, high school students are involved in the research in the context of this project. This offers a fascinating project for high school students, in which they combine their separate knowledge in biology, chemistry, physics and mathematics to conduct scientific research in a professional environment. The high school students will be supervised primarily by the graduate students involved in this project, which will give the UTMB students an excellent opportunity to teach at an institution with no regular undergraduate program. The international student exchange will provide opportunities for graduate students with different cultural backgrounds to learn the importance of wider vision in science, because each of the two groups is investigating the same subject by totally different and complementary approaches.
