TY - JOUR
T1 - Drug-RNA footprinting
AU - McPike, M. P.
AU - Goodisman, J.
AU - Dabrowiak, J. C.
N1 - Funding Information:
We thank P. N. Borer for the energy-minimized structure of the 176-mer and for helpful discussions pertaining to the research. This work was in part supported by a grant, GM32691, to P. N. Borer.
PY - 2001
Y1 - 2001
N2 - RNase I and RNase T1 can be used to obtain high-quality footprinting information for paromomycin binding to a 176-mer RNA from the packaging region of HIV-1 (LAI). Controls and scanning procedures are necessary for quantitation of autoradiographic data, so that footprinting plots showing cutting behavior as a function of drug concentration can be used to identify binding sites and regions of altered structure on the 176-mer. From the RNase I footprinting results the primary paromomycin binding sites on the 176-mer are on the main stem and on the stem of SL1, but noncontiguous sequences may be involved in the same binding event. Strong enhancements in cleavage with added drug are also observed, indicating drug-induced structural changes. Drug binding may cause linker regions between stem-loops of the 176-mer to change structure, possibly providing a site or sites for additional drug binding. Because drug binding changes the structure of the packaging region, which may alter its function, paromomycin analogs with enhanced specificity for HIV ψRNA have potential as a new class of agent for treating AIDS.
AB - RNase I and RNase T1 can be used to obtain high-quality footprinting information for paromomycin binding to a 176-mer RNA from the packaging region of HIV-1 (LAI). Controls and scanning procedures are necessary for quantitation of autoradiographic data, so that footprinting plots showing cutting behavior as a function of drug concentration can be used to identify binding sites and regions of altered structure on the 176-mer. From the RNase I footprinting results the primary paromomycin binding sites on the 176-mer are on the main stem and on the stem of SL1, but noncontiguous sequences may be involved in the same binding event. Strong enhancements in cleavage with added drug are also observed, indicating drug-induced structural changes. Drug binding may cause linker regions between stem-loops of the 176-mer to change structure, possibly providing a site or sites for additional drug binding. Because drug binding changes the structure of the packaging region, which may alter its function, paromomycin analogs with enhanced specificity for HIV ψRNA have potential as a new class of agent for treating AIDS.
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U2 - 10.1016/S0076-6879(01)40435-6
DO - 10.1016/S0076-6879(01)40435-6
M3 - Article
C2 - 11494862
AN - SCOPUS:0034890806
SN - 0076-6879
VL - 340
SP - 431
EP - 449
JO - Methods in enzymology
JF - Methods in enzymology
ER -