Physiologic and Transcriptomic Effects Triggered by Overexpression of Wild Type and Mutant DNA Topoisomerase I in Streptococcus pneumoniae
We obtained the topARA mutant that encodes a TopoI enzyme that shows reduced topoisomerase activity compared to the wild type gene (topAWT) and confers resistance to the antibiotic seconeolitsine. Alterations in the gene expression profile produced after overexpression of the wild type or the RA mutant of topA gene were studied. In addition, differential gene expression produced by the DNA gyrase inhibitor novobiocin (NOV) was also studied in cultures overexpressing each of the versions of TopA. Overall design: S. pneumoniae ?topAPZntopA strain was transformed with plasmid pLS1ROMtopAWT or pLS1ROMtopARA, where topA is under the control of a maltose-inducible promoter. Overnight cultures grown in medium with sucrose (noninducing S medium) were diluted 1000-fold in noninducing S medium or in sucrose+maltose medium (inducing SM medium) and grown to OD=0.4. The last 30 min of culture took place without NOV or with NOV 10xMIC.