Analysis of features contributing to activity of the single-stranded origin of Bacillus plasmid pBAA1.
The features which contribute to the activity of the single-stranded origin of the Bacillus plasmid pBAA1 were investigated. This origin is contained on a DNA fragment greater than 116 but less than 191 bases in size. There is the potential to form three stem-loop structures within this fragment. Comparison of the sequence of this origin from pBAA1 with the sequence of a homologous fragment from the Bacillus thuringiensis plasmid pGI2 indicates that both the structure and the relative positioning of the predicted stem-loops are important for origin activity. Deletion analysis suggests that it is the structure of stem-loop III which is important, because it can be replaced by a nonrelated dyad element without significant loss of origin activity. Three sequence motifs are conserved between the origins from pBAA1 and pGI2. Mutation of motif 1 leads to attenuation of single-stranded origin activity. A second motif (motif 3) shares significant homology with a group of single-strand initiation (ssi) sites found on plasmids isolated from Escherichia coli, suggesting that it also contributes to single-stranded origin activity. Our results also indicate that RNA polymerase is utilized to synthesize the RNA primer at the pBAA1 single-stranded origin and that this origin can function in both Bacillus subtilis and Staphylococcus aureus.
Seery
L
L
Department of Genetics, Trinity College, Dublin, Ireland.
Devine
K M
KM
D001412:Bacillus subtilis / Q000235:genetics* / Q000378:metabolism; D001413:Bacillus thuringiensis / Q000235:genetics; D001483:Base Sequence; D004252:DNA Mutational Analysis; D004261:DNA Replication; D004269:DNA, Bacterial / Q000096:biosynthesis / Q000235:genetics*; D004277:DNA, Single-Stranded; D012321:DNA-Directed RNA Polymerases / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D010957:Plasmids / Q000235:genetics*; D017384:Sequence Deletion; D013045:Species Specificity; D013329:Structure-Activity Relationship
D004269:DNA, Bacterial; D004277:DNA, Single-Stranded; D012321:DNA-Directed RNA Polymerases