The objectives of the proposed research are to determine the detailed structure [and to decipher the sequence-dependence of the structure] of double helical nucleic acids containing both RNA and DNA in the same duplex. Major emphasis will be placed on the sequence dependence of the structure of RNA:DNA hybrid duplexes and on covalent chimeric duplexes containing hybrid segments joined to DNA segments as well as hybrid segments joined to RNA segments. Such chimeric duplexes are extremely important in biology in such processes as the initiation of DNA synthesis, the continued replication of DNA, and the growth of retroviruses - yet little is known of their detailed structure and the importance of sequence on their function. We have recently demonstrated that many current ideas concerning the structure of hybrid duplexes and hybrid duplex junctions are very likely to be crystallographic artifacts. Hence all of the proposed structural studies will be carried out in solution using high-resolution NMR spectroscopy. These biologically important structures determined directly in solution will have widespread importance not only in modern molecular biology but will also generate new targets for specifically inhibiting the life-cycle of retroviruses.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
2R01GM042896-06A1
Application #
2181714
Study Section
Molecular and Cellular Biophysics Study Section (BBCA)
Project Start
1989-07-01
Project End
1998-04-30
Budget Start
1995-05-01
Budget End
1996-04-30
Support Year
6
Fiscal Year
1995
Total Cost
Indirect Cost
Name
University of Washington
Department
Chemistry
Type
Schools of Arts and Sciences
DUNS #
135646524
City
Seattle
State
WA
Country
United States
Zip Code
98195
Salazar, M; Fedoroff, O Y; Reid, B R (1996) Structure of chimeric duplex junctions: solution conformation of the retroviral Okazaki-like fragment r(ccca)d(AATGA).d(TCATTTGGG) from Moloney murine leukemia virus. Biochemistry 35:8126-35
Fedoroff OYu; Salazar, M; Reid, B R (1996) Structural variation among retroviral primer-DNA junctions: solution structure of the HIV-1 (-)-strand Okazaki fragment r(gcca)d(CTGC).d(GCAGTGGC). Biochemistry 35:11070-80
Zhu, L; Reid, B R (1995) An improved NOESY simulation program for partially relaxed spectra: BIRDER. J Magn Reson B 106:227-35
Zhu, L; Salazar, M; Reid, B R (1995) DNA duplexes flanked by hybrid duplexes: the solution structure of chimeric junctions in [r(cgcg)d(TATACGCG)]2. Biochemistry 34:2372-80
Wang, A C; Kennedy, M A; Reid, B R et al. (1994) A solid-state 2H NMR investigation of purine motion in a 12 base pair RNA duplex. J Magn Reson B 105:1-10
Salazar, M; Fedoroff OYu; Zhu, L et al. (1994) The solution structure of the r(gcg)d(TATACCC):d(GGGTATACGC) Okazaki fragment contains two distinct duplex morphologies connected by a junction. J Mol Biol 241:440-55
Fedoroff OYu; Salazar, M; Reid, B R (1993) Structure of a DNA:RNA hybrid duplex. Why RNase H does not cleave pure RNA. J Mol Biol 233:509-23
Salazar, M; Champoux, J J; Reid, B R (1993) Sugar conformations at hybrid duplex junctions in HIV-1 and Okazaki fragments. Biochemistry 32:739-44
Salazar, M; Fedoroff, O Y; Miller, J M et al. (1993) The DNA strand in DNA.RNA hybrid duplexes is neither B-form nor A-form in solution. Biochemistry 32:4207-15
Wang, A C; Kim, S G; Flynn, P F et al. (1992) Errors in RNA NOESY distance measurements in chimeric and hybrid duplexes: differences in RNA and DNA proton relaxation. Biochemistry 31:3940-6

Showing the most recent 10 out of 11 publications