The major goal of the proposed research is a detailed underestanding of the forces primarily responsible for determining the long-range structure of DNA. Such an understanding will aid in analyzing the means by which DNA is organized into compact structures (e.g., chromosomes and viral particles) as well as the mechanisms by which DNA structure is altered by small ligands (e.g., intercalating drugs), DNA binding proteins, and disease processes which later its chemistry) (e.g., base removal, single-strand nicking, etc.). A knowledge of such mechanisms is essential for the broader understanding of the regulation of gene expression. The primary aim of this application is to identify those features of DNA which contribute to its long-range order. This examination will be carried out in a systematic fashion by varying the DNA sequence, the integrity of its phosphodiester backbone, etc. My experimental approach comprises two parts; namely, (1) the use of recombinant DNA cloning methodologies to produce a wide variety of DNA molecules of precisely defined sequence and length, and (2) the use of two independent, self-consistent experimental methods for measuring the flexibility of DNA. The first method consists of the measurement of rotational relaxation times of DNA molecules of interest by following the field-free decay of birefringence (TEB). The second method consists of the measurement of the rates of formation of small circles (catalyzed by T4 DNA ligase).
The second aim of this application is to further characterize the electrostatic contribution to the rigidity of DNA at low ionic strengths.
The third aim of this application is to characterize the pathway of interconversion between B-form and Z-form DNA by performing kinetic TEB measurements, utilizing a prototype stopped-flow TEB instrument.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM028293-06
Application #
3275577
Study Section
Biophysics and Biophysical Chemistry A Study Section (BBCA)
Project Start
1980-07-01
Project End
1986-06-30
Budget Start
1985-07-01
Budget End
1986-06-30
Support Year
6
Fiscal Year
1985
Total Cost
Indirect Cost
Name
University of Colorado Denver
Department
Type
Schools of Medicine
DUNS #
065391526
City
Aurora
State
CO
Country
United States
Zip Code
80045
Hagerman, K R; Hagerman, P J (1996) Helix rigidity of DNA: the meroduplex as an experimental paradigm. J Mol Biol 260:207-23
Mills, J B; Cooper, J P; Hagerman, P J (1994) Electrophoretic evidence that single-stranded regions of one or more nucleotides dramatically increase the flexibility of DNA. Biochemistry 33:1797-803
Hagerman, P J (1992) Straightening out the bends in curved DNA. Biochim Biophys Acta 1131:125-32
Taylor, W H; Hagerman, P J (1990) Application of the method of phage T4 DNA ligase-catalyzed ring-closure to the study of DNA structure. II. NaCl-dependence of DNA flexibility and helical repeat. J Mol Biol 212:363-76
Hagerman, P J (1990) Pyrimidine 5-methyl groups influence the magnitude of DNA curvature. Biochemistry 29:1980-3
Hagerman, P J (1990) Sequence-directed curvature of DNA. Annu Rev Biochem 59:755-81
Hagerman, P J; Ramadevi, V A (1990) Application of the method of phage T4 DNA ligase-catalyzed ring-closure to the study of DNA structure. I. Computational analysis. J Mol Biol 212:351-62
Hagerman, P J (1988) Flexibility of DNA. Annu Rev Biophys Biophys Chem 17:265-86
Taylor, W H; Hagerman, P J (1987) A general method for cloning DNA fragments in multiple copies. Gene 53:139-44
Hagerman, P J (1986) Sequence-directed curvature of DNA. Nature 321:449-50

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