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Thr729 mutants are catalytically active in vitro. A total of 120 pmol of purified hTop1p, hTop1Thr729Ala, hTop1Thr729Pro, hTop1Thr729Lys and hTop1Thr729Glu were serially 10-fold diluted and incubated in DNA relaxation assays with negatively supercoiled plasmid DNA. Following incubation at 37°C for 30 min, the reaction products were resolved in agarose gels and visualized after staining with ethidium bromide. C indicates no enzyme control.
