StpA protein from Escherichia coli condenses supercoiled DNA in preference to linear DNA and protects it from digestion by DNase I and EcoKI.
The nucleoid-associated protein, StpA, of Escherichia coli binds non-specifically to double-stranded DNA (dsDNA) and apparently forms bridges between adjacent segments of the DNA. Such a coating of protein on the DNA would be expected to hinder the action of nucleases. We demonstrate that StpA binding hinders dsDNA cleavage by both the non-specific endonuclease, DNase I, and by the site-specific type I restriction endonuclease, EcoKI. It requires approximately one StpA molecule per 250-300 bp of supercoiled DNA and approximately one StpA molecule per 60-100 bp on linear DNA for strong inhibition of the nucleases. These results support the role of StpA as a nucleoid-structuring protein which binds DNA segments together. The inhibition of EcoKI, which cleaves DNA at a site remote from its initial target sequence after extensive DNA translocation driven by ATP hydrolysis, suggests that these enzymes would be unable to function on chromosomal DNA even during times of DNA damage when potentially lethal, unmodified target sites occur on the chromosome. This supports a role for nucleoid-associated proteins in restriction alleviation during times of cell stress.
Keatch
S A
SA
School of Chemistry, The King's Buildings, The University of Edinburgh, Edinburgh EH9 3JJ, UK.
Leonard
P G
PG
Ladbury
J E
JE
Dryden
D T F
DT
D000255:Adenosine Triphosphate / Q000378:metabolism; D004262:DNA Restriction Enzymes / Q000378:metabolism*; D004269:DNA, Bacterial / Q000737:chemistry / Q000378:metabolism*; D004278:DNA, Superhelical / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000378:metabolism*; D003850:Deoxyribonuclease I / Q000378:metabolism*; D029968:Escherichia coli Proteins / Q000378:metabolism*; D018832:Molecular Chaperones / Q000378:metabolism*
D004269:DNA, Bacterial; D004278:DNA, Superhelical; D004268:DNA-Binding Proteins; D029968:Escherichia coli Proteins; D018832:Molecular Chaperones; C090104:StpA protein, E coli; D000255:Adenosine Triphosphate; D004262:DNA Restriction Enzymes; C089629:endodeoxyribonuclease EcoKI; D003850:Deoxyribonuclease I