Plasmid transcriptional repressor CopG oligomerises to render helical superstructures unbound and in complexes with oligonucleotides

CopG is a 45 amino acid residue transcriptional repressor involved in the c opy number control of the streptococcal plasmid pMV158. To do so, it binds to a DNA operator that contains a 13 bp pseudosymmetric DNA element. Bindin g of CopG to its operator results in repression, at the transcriptional lev el, of its own synthesis and that of the initiator of replication protein, RepB. Biochemical experiments have shown that CopG co-operatively associate s to its target DNA at low protein:DNA ratios, completely protecting four h elical turns on the same face of the double helix in both directions from t he inverted repeat that constitutes the CopG primary target. This has been correlated with a CopG-mediated DNA bend of about 100 degrees Here, we show that binding of CopG to DNA fragments containing the inverted repeat just at one end led to nucleation of the protein initiating from the inverted re peat. Nucleation extended to the entire fragment, with CopG-DNA contacts oc curring on the same face of the DNA helix. The protein, the prototype for a family of homologous plasmid repressors, displays a homodimeric ribbon-hel ix-helix arrangement. It polymerises within the unbound crystal to render a continuous right-handed protein superhelix of homodimers, around which a b ound double-stranded (ds) DNA could wrap. We have solved the crystal struct ure of CopG in complex with a 22 bp dsDNA oligonucleotide encompassing the cognate pseudosymmetric element. In the crystal, one protein tetramer binds at one face of the DNA with two parallel P-ribbons inserted into the major groove. The DNA is bent about 50 degrees under compression of both major a nd minor grooves. A continuous right-handed complex helix made up mainly by protein-protein and some protein-DNA interactions is observed. The protein -protein inter actions involve regions similar to those observed in the oli gomerisation of the native crystals and those employed to set up the functi onal tetramer. A previously solved complex structure of the protein with a 19 bp dsDNA had unveiled a left-handed helical superstructure just made up by DNA interactions. (C) 2001 Academic Press.