Electron microscopic mapping of proteins bound to herpes simplex virus DNA.
Exonuclease digestion experiments have suggested that there is a protein(s) bound close to one or both ends of herpes simplex virus-1 (HSV) DNA. The existence of such bound proteins has been positively demonstrated and their positions on the HSV genome determined by application of a newly developed method for electron microscopic mapping of proteins bound to nucleic acids. Purified HSV DNA was treated with dinitrofluorobenzene under conditions that covalently attach the dinitrophenyl (DNP) group to the proteins in a protein-nucleic acid complex. The HSV DNA-protein-(DNP)n complex was treated with rabbit anti-DNP IgG, and, in some cases, additionally treated with monovalent Fab fragments of goat anti-rabbit IgG, and mounted for examination in the electron microscope. Electron opaque dots representing the protein-(DNP)n-(IgG)m complex were seen on the HSV DNA. Direct measurements of the positions of the protein, as well as partial denaturation mapping, indicate that there are four positions for protein bound to HSV DNA: two near but not at the two ends and two at sites corresponding to the internal inverted repeats of the ends. These results suggest that there is a specific protein binding sequence within the direct terminal repeat of HSV DNA. The previous observation that HSV DNA is more sensitive to digestion by a 3' than by a 5' exonuclease then indicates that the bound protein(s) is more intimately associated with one strand of the specific sequence than with the complementary strand.
D001665:Binding Sites; D004279:DNA, Viral / Q000032:analysis*; D003851:Deoxyribonucleases; D004139:Dinitrofluorobenzene; D005092:Exonucleases; D008854:Microscopy, Electron; D009690:Nucleic Acid Conformation; D011485:Protein Binding; D011487:Protein Conformation; D018139:Simplexvirus / Q000032:analysis*; D014764:Viral Proteins / Q000032:analysis*
D004279:DNA, Viral; D014764:Viral Proteins; D004139:Dinitrofluorobenzene; D003851:Deoxyribonucleases; D005092:Exonucleases