Abstract
Three regions showing abnormal electrophoretic mobility, which is an indication of the existence of bends in DNA, have been observed in the DNA of plasmid pLS1. These loci have been characterized by assays designed to detect sequence-directed bending in DNA (temperature-dependence migration and two dimensional electrophoresis). The first region (locus B-1) was located within a fragment that contains a proposed inhibitor countertranscribed RNA (RNAII). The second locus (B-2) contains the plasmid plus origin of replication and the third region (locus B-3) was located in the vicinity of a putative antisense RNA (RNAI) of unknown function. The centres of the first two bent DNA regions were located by circular permutation assays at nucleotides 882 (locus B-1) and 634 (locus B-2). The bend centre of locus B-1 was found to be upstream of the promoter for the putative antisense RNAII. The centre of curvature in locus B-2 was located in the vicinity of the putative promoter of the replication proteins RepA and RepB and of a sequence that has three 11-bp direct repeats. The DNA sequence at this region showed the existence of A.T tracts, with an internal repeat of 10 to 11 base pairs, for five helix turns. A complex curvature in the DNA of pLS1 at locus B-2 that may have a regulatory role in plasmid replication is postulated.