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. 2018 Dec 19;47(3):1416–1427. doi: 10.1093/nar/gky1271

Figure 1.

Figure 1.

Experimental approaches and validation. (A) Schematic of experimental approaches to map TopoI functional sites. Imipramine treated (IT) and D108A mutant mediated protein-DNA covalent adducts were immune-precipated using anti-TopoI antibodies and sequenced. (B) Overall occupancy of TopoI is determined by formaldehyde crosslinking of TopoI and DNA (FC). TopoI linked to DNA was immune-precipitated using anti-TopoI antibodies. (C) Dot-blots showing Protein-DNA covalent complexes isolated from MsTopoI D108A overexpressing M. smegmatis (mc2 155) cells and (E) imipramine treated M. smegmatis cells probed with TopoI antibodies. (D) qPCR showing increased recA expression in MsTopoI D108A overexpressing M. smegmatis cells and (F) imipramine treated M. smegmatis cells. The data represented is mean ± SD from the three independent experiments. UT = untreated, IT = imipramine treated.