title: An electron microscopic method for the mapping of proteins attached to nucleic acids.
abstract: An electron microscopic method for demonstrating the presence of and mapping the positions of proteins specifically bound to nucleic acids is described. The nucleic acid-protein complex is treated with dinitrofluorobenzene under conditions such that dinitrophenyl (DNP) groups are attached to nucleophilic groups on the protein, with only a low level of random attachment to the nuclei acid. This product is treated with rabbit anti-DNP IgG. The position of the protein-(DNP)n(IgG)m complex on the nucleic acid strand can be observed by electron microscopy by protein free spreading methods and, in many cases, by cytochrome-c spreading. If necessary for visualization by the latter method, the size of the labeled region can be increased by treatment with goat anti-rabbit IgG. High efficiency of electron microscopic labeling is achieved. Examples studied are: the adenovirus-2 DNA terminal protein, a protein covalently bound to SV40 DNA, DNA polymerase I bound to DNA, E. coli RNA polymerase bound to T7 DNA, and proteins UV crosslinked to avian sarcoma virus RNA.
mesh_terms: D000260:Adenoviruses, Human; D001358:Avian Sarcoma Viruses / Q000528:radiation effects; D001665:Binding Sites; D003090:Coliphages / Q000201:enzymology; D004279:DNA, Viral / Q000378:metabolism*; D012321:DNA-Directed RNA Polymerases / Q000378:metabolism; D007158:Immunologic Techniques; D008854:Microscopy, Electron / Q000379:methods*; D013539:Simian virus 40; D014466:Ultraviolet Rays; D014764:Viral Proteins / Q000378:metabolism*
chemical_list: D004279:DNA, Viral; D014764:Viral Proteins; D012321:DNA-Directed RNA Polymerases