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.

National Institute of Health (NIH)
National Institute of General Medical Sciences (NIGMS)
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Macromolecular Structure and Function B Study Section (MSFB)
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Preusch, Peter C
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Mayo Clinic, Rochester
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Peters, Justin P; Kowal, Ewa A; Pallan, Pradeep S et al. (2018) Comparative analysis of inosine-substituted duplex DNA by circular dichroism and X-ray crystallography. J Biomol Struct Dyn 36:2753-2772
Murugesapillai, Divakaran; Bouaziz, Serge; Maher, L James et al. (2017) Accurate nanoscale flexibility measurement of DNA and DNA-protein complexes by atomic force microscopy in liquid. Nanoscale 9:11327-11337
Uchida, Akira; Murugesapillai, Divakaran; Kastner, Markus et al. (2017) Unexpected sequences and structures of mtDNA required for efficient transcription from the first heavy-strand promoter. Elife 6:
Murugesapillai, Divakaran; McCauley, Micah J; Maher 3rd, L James et al. (2017) Single-molecule studies of high-mobility group B architectural DNA bending proteins. Biophys Rev 9:17-40
Mogil, Lauren S; Becker, Nicole A; Maher 3rd, L James (2016) Supercoiling Effects on Short-Range DNA Looping in E. coli. PLoS One 11:e0165306
Becker, Nicole A; Maher 3rd, L James (2015) High-resolution mapping of architectural DNA binding protein facilitation of a DNA repression loop in Escherichia coli. Proc Natl Acad Sci U S A 112:7177-82
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
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
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
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 107:448-459

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