Abstract
A site-specific DNA-binding protein was purified from Borrelia burgdorferi cytoplasmic extracts, and determined to be a member of the highly conserved SpoVG family. This is the first time a function has been attributed to any of these ubiquitous bacterial proteins. Further investigations into SpoVG orthologues indicated that the Staphylococcus aureus protein also binds DNA, but interacts preferentially with a distinct nucleic acid sequence. Site-directed mutagenesis and domain swapping between the S. aureus and B. burgdorferi proteins identified that a 6-residue stretch of the SpoVG α-helix contributes to DNA sequence specificity. Two additional, highly conserved amino acid residues on an adjacent β-sheet are essential for DNA-binding, apparently by contacts with the DNA phosphate backbone. Results of these studies thus identified a novel family of bacterial DNA-binding proteins, developed a model of SpoVG-DNA interactions, and provide direction for future functional studies on these wide-spread proteins.
Highlights
To be successful, single-celled organisms must efficiently and rapidly adapt to changing conditions
In our studies of the VlsE antigenic variation system of Borrelia burgdorferi, the causative agent of Lyme disease [1,2], we discovered that these bacteria produce a cytoplasmic protein which binds a DNA site within the vlsE open reading frame
DNAs flanking those 70 bp did not compete for protein binding (Fig. 1B, lanes 3 and 5). These results indicate that the borrelial protein binds a DNA sequence of approximately 70 bp (X on Fig. 1A), and that neither of the repeat regions flanking the recombination site is involved in protein binding
Summary
Single-celled organisms must efficiently and rapidly adapt to changing conditions. This is often accomplished through exquisite regulatory networks involving numerous, dynamic trans-acting factors. Prokaryotic proteins that bind to nucleic acids govern virtually every cellular process, including nucleoid organization, transcription, translation, and DNA replication, modification, repair, and recombination. Most DNA-binding proteins are poorly characterized, and it is likely that many more await discovery. In our studies of the VlsE antigenic variation system of Borrelia burgdorferi, the causative agent of Lyme disease [1,2], we discovered that these bacteria produce a cytoplasmic protein which binds a DNA site within the vlsE open reading frame. Until our discovery, the biochemistry of SpoVG remained a mystery
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