Figure 2.
Expression, purification, and activity of R1Bm EN protein. (a) R1Bm EN protein and its mutant versions E40A and D186A were expressed from the phage T7 promoter. The proteins were purified by nickel chelate chromatography and run on SDS/PAGE; the predicted Mr is 29,000. The unusual electrophoretic mobility of the D186A mutant was observed for two independent isolates. (b) The activity of the R1Bm EN protein was optimized and tested on two plasmid substrates, pBluescript (KS−) vector and pB109. The positions of open circle (oc), linear, and supercoiled (sc) plasmid controls are indicated on the right. Note that the mutant proteins are inactive. MW, molecular weight standards.