N-terminal amino acid residues mediate protein-protein interactions between DNA-bound alpha/ss-type small, acid-soluble spore proteins from Bacillus species

The binding of alpha/beta -type small, acid-soluble spore proteins (SASP) t o DNA of spores of Bacillus species is the major determinant of DNA resista nce to a variety of damaging treatments. The primary sequence of alpha/beta -type SASP is highly conserved; however, the N-terminal third of these pro teins is less well conserved than the C-terminal two-thirds. To determine t he functional importance of residues in the N-terminal region of alpha/beta -type SASP, variants of SspC (a minor alpha/beta -type SASP from Bacillus subtilis) with modified N termini were generated and their structural and D NA binding properties studied in vitro and in vivo. SspC variants with dele tions of up to 14 residues (similar to 20% of SspC residues) were able to b ind DNA in vitro and adopted similar conformations when bound to DNA, as de termined by circular dichroism spectroscopy and protein-protein cross-linki ng. Progressive deletion of up to 11 N-terminal residues resulted in protei ns with progressively lower DNA binding affinity. However, SspC(Delta 14) ( in which 14 N-terminal residues have been deleted) showed significantly hig her affinity for DNA than the larger proteins, SspC(Delta 10) and SspC(Delt a 11). The affinity of these proteins for DNA was shown to be largely depen dent upon the charge of the first few N-terminal residues. These results ar e interpreted in the context of a model for DNA-dependent alpha/beta -type SASP protein-protein interaction involving the N-terminal regions of these proteins.