| Summary: | 碩士 === 國立清華大學 === 生命科學系 === 88 === We have determined the solution structure of DNA/RNA hybrid chimeric duplex [d(CGC)r(aaa)d(TTTGCG)]2 using high resolution NMR spectroscopy, simulated annealing and restraint molecular dynamics. The solution structure of this hybrid duplex differs from its DNA duplex analog d(CGCAAATTTGCG)2 solved by X-ray crystallography (Edwards, et al 1992) and its RNA duplex analog r(cgcaaauuugcg)2 solved by NMR (Conte et al., 1997) as well. However, the overall structure is closer to a typical B-form DNA with a narrow minor groove width of about 6 A in the central hybrid segment. Long-lived water molecules with water correlation time tc longer than 0.3 ns were observed around the H2 and H1' protons of three RNA adenine residues as well as the methyl group of the thymine at the 7 position by combination of the laboratory frame nuclear Overhauser effect spectroscopy (NOESY) and the rotating frame NOE spectroscopy (ROESY). The unusual hydration pattern of the 7T methyl group in the major groove has not been observed in the previous NMR studies. The sugar ribose of 7T in the RNA-DNA hybrid junction was found to adopt a O4'-endo conformation, while other DNA residues, including 3C in the DNA-RNA hybrid junction, adopted C1'-exo or C2'-endo conformations. X-ray crystallography study of RNA duplex hydration also suggested a hydration network bridged via the C2'-OH of the RNA ribose with a C3'-endo conformation (Egli et al., 1996). The special sugar conformation of the 7T together with the additional C2'-OH of the 5'-adjacent RNA adenine may provide a hydraulic environment and thus showed a different hydration pattern from the other two thymine residues, 8T and 9T. The exchange rate of the RNA C2'-OH was found to be ~ 5-20 s-1. This slow exchange rate may due to the narrow minor groove width which restricts the dynamic motion of the hydroxyl protons. We further examined the structural role of the C2'-OH of the RNA residues by a 2'-O-methylated analog [d(CGC)r(amamam)d(TTTGCG)]2. The structure was not altered significantly, but the hydration of the 7T methyl decreased and it became merely observable in the minor groove. These distinct structural features and hydration patterns reveal the importance of the C2'-OH in the DNA/RNA hybrid structure and therefore provide a potential target in molecular recognition.
|
|---|
