Alteration of the curved helical structure located in the upstream region of the β-lactamase promoter of plasmid pUC 19 and its effect on transcription

The region preceding the β-lactamase promoter of Escherichia coli plasmid pUC19 has a curved DNA (bent DNA) structure. The center of the curvature was revealed to exist around nucleotide position 2580 of the plasmid, which is just beside RNA polymerase binding region. It was indicated that the identified region is curved even at 60° C. The gross geometry of the curvature was altered by inserting synthetic double-stranded oligonucleotides between positions 2585 and 2586. Effect of the alteration on strength of the promoter was not detectedin vitro. However, in vivo analyses showed that the promoter strength is apparently dependent, in part, on the gross geometry of the curvature. Insertions of 4 and 16 bp, both of which altered the gross geometry of the curvature greatly, caused considerable reductions of in vivo level of β-lactamase mRNA. in vivo, overall three-dimensional structure of the region covering the promoter and the curvature seems to play some significant role in transcription of the gene.