Our laboratory is interested in understanding the physical origin and biological management of DNA stiffness. It has long been appreciated that duplex DNA is among the stiffest of all natural polymers. Remarkably, the origin of DNA stiffness remains obscure after 50 years of study. A better understanding of the balance of forces within DNA would reveal what forces are overcome by DNA bending proteins such as histone octamers, and could allow control of DNA stiffness for nanomaterials and nanodevices. How living cells manage the stiff DNA polymer is also not understood in mechanistic detail. The bending and twisting flexibility of DNA within cells appears to be higher than in solution. Why? We have been studying sequence non-specific HMGB DNA bending proteins and other "architectural" proteins as models for understanding one mechanism by which DNA might be endowed with enhanced apparent flexibility in cells. Understanding and harnessing the properties of these proteins could have applications in artificial gene control by targeting the proteins to modulate the stability of DNA loops required for transcriptional regulation. During the previous funding period we made important progress toward understanding the origin and management of DNA stiffness. We now propose four aims to continue this fundamental research.
Aim 1 will determine the relationship between DNA charge density, base stacking and DNA stiffness.
Aim 2 will measure effects of HMGB protein binding on the flexibility and structure of DNA and chromatin in vitro.
Aim 3 will improve our understanding of the basis for DNA looping enhancement by architectural proteins in E. coli. Finally, Aim 4 will measure the effects of HMGB proteins on DNA looping in yeast.

Public Health Relevance

DNA molecules contain the information code for all living things. This information is contained in very long double-helix DNA molecules. These molecules are thread-like when considered at a distance, but are rod-like from the perspective of the proteins that must bind and read DNA. This proposal for renewed funding will allow our collaborative research group of molecular biologists, biochemists, and physicists to continue our productive projects to understand why DNA is stiff and rod-like locally, and how a special group of proteins called architectural proteins increase the flexibility of DNA by causing bends and kinks.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM075965-07
Application #
8473878
Study Section
Macromolecular Structure and Function B Study Section (MSFB)
Program Officer
Preusch, Peter C
Project Start
2006-09-25
Project End
2015-05-31
Budget Start
2013-06-01
Budget End
2014-05-31
Support Year
7
Fiscal Year
2013
Total Cost
$342,509
Indirect Cost
$82,177
Name
Mayo Clinic, Rochester
Department
Type
DUNS #
006471700
City
Rochester
State
MN
Country
United States
Zip Code
55905
Peters, Justin P; Mogil, Lauren S; McCauley, Micah J et al. (2014) Mechanical properties of base-modified DNA are not strictly determined by base stacking or electrostatic interactions. Biophys J 107:448-59
Stellwagen, Nancy C; Peters, Justin P; Dong, Qian et al. (2014) The free solution mobility of DNA and other analytes varies as the logarithm of the fractional negative charge. Electrophoresis 35:1855-63
Becker, Nicole A; Greiner, Alexander M; Peters, Justin P et al. (2014) Bacterial promoter repression by DNA looping without protein-protein binding competition. Nucleic Acids Res 42:5495-504
Murugesapillai, Divakaran; McCauley, Micah J; Huo, Ran et al. (2014) DNA bridging and looping by HMO1 provides a mechanism for stabilizing nucleosome-free chromatin. Nucleic Acids Res 42:8996-9004
Coats, Julie E; Lin, Yuyen; Rueter, Emily et al. (2013) Single-molecule FRET analysis of DNA binding and bending by yeast HMGB protein Nhp6A. Nucleic Acids Res 41:1372-81
Stellwagen, Earle; Peters, Justin P; Maher 3rd, L James et al. (2013) DNA A-tracts are not curved in solutions containing high concentrations of monovalent cations. Biochemistry 52:4138-48
Becker, Nicole A; Peters, Justin P; Maher 3rd, L James et al. (2013) Mechanism of promoter repression by Lac repressor-DNA loops. Nucleic Acids Res 41:156-66
McCauley, Micah J; Rueter, Emily M; Rouzina, Ioulia et al. (2013) Single-molecule kinetics reveal microscopic mechanism by which High-Mobility Group B proteins alter DNA flexibility. Nucleic Acids Res 41:167-81
Zhang, Jingyun; McCauley, Micah J; Maher 3rd, L James et al. (2012) Basic N-terminus of yeast Nhp6A regulates the mechanism of its DNA flexibility enhancement. J Mol Biol 416:10-20
McCauley, Micah J; Williams, Mark C (2011) Measuring DNA-protein binding affinity on a single molecule using optical tweezers. Methods Mol Biol 749:305-15

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