Abstract

1. Polynucleotide Structures. (a) Structures Stabilized by C.C+ Base pairs. DNA sequencing work by others has led to the possibility that C-rich non-coding regions of DNA may be important in DNA replication and gene recombination. DNA sequences such as poly(d(CnTm)) and poly(d(CnAm)) with varying n and m will be synthesized and studied as a function of pH. Sequences with n greater than m should be able to form novel loop-out structures, with unpaired T or A bases being excluded from a helix of stacked, hemi- protonated C.C+ base pairs. Such structures will be studied by photochemical analyses as well as by circular dichroism (CD) spectroscopy. Those sequences with equal numbers of A and T bases will also be mixed in solution to determine limitations on the number of adjacent, stacked C.C+ base pairs that can coexist with A.T base pairs in a double helix under physiological conditions. (b) DNA.RNA hybrids. Hybrids have been the least studied area of polynucleotide structure, even though their properties are important in the control of transcription and replication. Seven new hybrids will be synthesized by a procedure of transcription form I- containing polymers, developed by Dr. Ratliff. In addition to investigating their solution conformations by CD spectroscopy in the near UV, they will be studied by vacuum UV CD. Selected hybrids and sequences that form stable C.C+ pairs will also be used for fiber X-ray diffraction studies. 2. Polynucleotide-Protein Structures. The gene 5 protein (G5P) of fd phage is a model single- strand DNA binding protein whose binding site has yet to be adequately characterized. Properties of this protein will be explored in three ways. (a) The two stoichoimetric binding modes of the fd G5P and related Ike G5P will be studied by CD titrations of synthetic DNA sequences. (b) 13C NMR measurements on reductively methylated G5P, combined with sequencing, will be used to determine which of the 6 lysyl residues of the protein are specifically involved in the DNA-binding function. (c) Neutron scattering will be used to study the solution structures of the two proteins and the complexes they form in different stoichiometric modes.

  Funding Agency

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM019060-16
Application #
3269495
Study Section
Biophysics and Biophysical Chemistry A Study Section (BBCA)
Project Start
1978-08-01
Project End
1992-07-31
Budget Start
1988-08-01
Budget End
1989-07-31
Support Year
16
Fiscal Year
1988
Total Cost
Indirect Cost

  Institution

Name
University of Texas-Dallas
Department
Type
Schools of Arts and Sciences
DUNS #
City
Richardson
State
TX
Country
United States
Zip Code
75080

  Publications

Antao, V P; Gray, D M (1993) CD spectral comparisons of the acid-induced structures of poly[d(A)], poly[r(A)], poly[d(C)], and poly[r(C)] J Biomol Struct Dyn 10:819-39
Powell, M D; Gray, D M (1993) Characterization of the Pf3 single-strand DNA binding protein by circular dichroism spectroscopy. Biochemistry 32:12538-47
Peterson, C A; Gray, D M; Gray Jr, H B et al. (1993) Evidence for a salt-induced conformational transition in UV-irradiated superhelical PM2 DNA. Biochim Biophys Acta 1216:265-72
Clack, B A; Gray, D M (1992) Flow linear dichroism spectra of four filamentous bacteriophages: DNA and coat protein contributions. Biopolymers 32:795-810
Gray, D M; Ratliff, R L; Vaughan, M R (1992) Circular dichroism spectroscopy of DNA. Methods Enzymol 211:389-406
Johnson, K H; Gray, D M (1992) Analysis of an RNA pseudoknot structure by CD spectroscopy. J Biomol Struct Dyn 9:733-45
Johnson, K H; Gray, D M (1991) A method for estimating the nearest neighbor base-pair content of RNAs using CD and absorption spectroscopy. Biopolymers 31:373-84
Johnson, K H; Gray, D M (1991) An estimate of the nearest neighbor base-pair content of 5S RNA using CD and absorption spectroscopy. Biopolymers 31:385-95
Johnson, K H; Gray, D M; Sutherland, J C (1991) Vacuum UV CD spectra of homopolymer duplexes and triplexes containing A.T or A.U base pairs. Nucleic Acids Res 19:2275-80
Johnson, K H; Gray, D M; Morris, P A et al. (1990) A.U and G.C base pairs in synthetic RNAS have characteristic vacuum UV CD bands. Biopolymers 29:325-33

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