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AcrIIA22 nicks supercoiled plasmids in vivo and in vitro.

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posted on 13.10.2021, 17:49 by Kevin J. Forsberg, Danica T. Schmidtke, Rachel Werther, Ruben V. Uribe, Deanna Hausman, Morten O. A. Sommer, Barry L. Stoddard, Brett K. Kaiser, Harmit S. Malik

(A) Gel electrophoresis of plasmids purified from overnight E. coli cultures expressing either acrIIA22, or a null mutant with an early stop codon, or neither. Compared to the null mutant, more plasmid runs in a slowly migrating, OC conformation rather than SC plasmid with the wild-type acrIIA22 allele, suggesting that acrIIA22 may impact plasmid topology. %SC indicates the percentage of DNA in the SC form for each sample. (B) N-terminally His6-tagged AcrIIA22 nicks supercoiled plasmids in vitro. (C) C-terminally twin-strep-tagged AcrIIA22 nicks supercoiled plasmids in vitro with higher specific activity than shown in panel B (compare protein concentrations). Original, uncropped versions of images depicted in figure may be found in the Supporting information file, S1 Raw Images. OC, open-circle; SC, supercoiled.