title: Altered DNA conformations detected by mung bean nuclease occur in promoter and terminator regions of supercoiled pBR322 DNA.
abstract: Mung bean nuclease was used to probe for recognizable DNA unwinding and unpairing in the plasmid pBR322. In negatively supercoiled DNA, but not relaxed DNA, cleavages occurred preferentially in non-coding regions of the genome. The types of nucleotide sequences cleaved and which non-coding regions were cleaved depended upon environmental conditions. At 37 degrees C, cleavages occurred in an 84 bp A+T-rich sequence in the terminator region of the ampicillin-resistance gene. Recognition is likely based on a novel DNA conformation which occurs in the longest, most dA+dT-rich region of pBR322. In the presence of 1 mM Mg2+, cleavages occurred in inverted repeated sequences in the promoter regions of the RNA primer for DNA replication and ampicillin- and tetracycline-resistance genes as well as the terminator of RNA-1. Potential loops of hairpin (cruciform) structures were cleaved. At 27 degrees C, cleavages occurred near a promoter activated by cAMP receptor protein in vitro and in the 3' non-coding region of the tetracycline-resistance gene. Thus, in supercoiled pBR322 DNA, recognizable DNA unwinding and unpairing occurs preferentially in regulatory regions for transcription and DNA replication.
mesh_terms: D001483:Base Sequence; D004262:DNA Restriction Enzymes; D004278:DNA, Superhelical / Q000235:genetics*; D004720:Endonucleases / Q000378:metabolism*; D004926:Escherichia coli / Q000235:genetics; D005809:Genes, Regulator; D007700:Kinetics; D009690:Nucleic Acid Conformation; D010944:Plants / Q000201:enzymology; D010957:Plasmids; D011401:Promoter Regions, Genetic; D015719:Single-Strand Specific DNA and RNA Endonucleases; D013379:Substrate Specificity; D013728:Terminator Regions, Genetic
chemical_list: D004278:DNA, Superhelical; D004720:Endonucleases; D004262:DNA Restriction Enzymes; D015719:Single-Strand Specific DNA and RNA Endonucleases