Interaction between DNA and alpha/beta-type small, acid-soluble spore proteins: a new class of DNA-binding protein.
DNA in spores of Bacillus and Clostridium species is associated with small, acid-soluble proteins (SASP) of the alpha/beta type; the presence of these proteins is a major factor in causing spore resistance to UV light, alpha/beta-type SASP did not bind to single-stranded DNA, single- or double-stranded RNA, or DNA-RNA hybrids in vitro. However, these proteins bound a variety of double-stranded DNAs and conferred protection against DNase cleavage. The binding of alpha/beta-type SASP to DNA saturated at a protein/DNA ratio (wt/wt) of 4:1 to 5:1, which is approximately 1 SASP per 4 bp. alpha/beta-type SASP-DNA interaction did not require divalent cations, was independent of pH between 6 and 8, and, for some SASP-DNA pairs, was relatively insensitive to salt up to 0.3 M. The relative affinity of alpha/beta-type SASP for different DNAs was poly(dG).poly(dC) greater than poly(dG-dC).poly(dG-dC) greater than plasmid pUC19 greater than poly(dA-dT).poly(dA-dT), with poly(dA).poly(dT) giving no detectable binding. This order in alpha/beta-type SASP-DNA affinities parallels the facility with which the DNAs adopt an A-like conformation, the conformation in alpha/beta-type SASP-DNA complexes. An oligo(dG).oligo(dC) of 12 bp was bound by alpha/beta-type SASP. While a 26-bp oligo(dG).oligo(dC) bound more tightly than the 12-mer, there was no significant increase in affinity for alpha/beta-type SASP with further increase in size of oligo(dG).oligo(dC). In contrast, binding of alpha/beta-type SASP to oligo(dA-dT).oligo(dA-dT) was minimal up to at least a 70-mer, and binding to poly(dA-dT).poly(dA-dT) was very cooperative. In addition to blocking DNase digestion, binding of alpha/beta-type SASP to DNA blocked (i) cleavage of the DNA backbone by hydroxyl radicals and orthophenanthroline-Cu2+, (ii) DNA cleavage by restriction enzymes, in particular those with specificity for GC-rich sequences; and (iii) in vitro transcription of some but not all genes. However, methylation of dG residues by dimethyl sulfate was not affected by alpha/beta-type SASP binding.
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Department of Biochemistry, University of Connecticut Health Center, Farmington 06030.
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Setlow
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D001426:Bacterial Proteins / Q000737:chemistry / Q000378:metabolism* / Q000494:pharmacology; D001667:Binding, Competitive; D004262:DNA Restriction Enzymes / Q000378:metabolism; D004269:DNA, Bacterial / Q000378:metabolism*; D004277:DNA, Single-Stranded / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000378:metabolism*; D003851:Deoxyribonucleases / Q000378:metabolism; D015964:Gene Expression Regulation, Bacterial / Q000187:drug effects; D010957:Plasmids; D012808:Sigma Factor; D012995:Solubility; D013171:Spores, Bacterial; D014157:Transcription Factors; D014158:Transcription, Genetic
D001426:Bacterial Proteins; D004269:DNA, Bacterial; D004277:DNA, Single-Stranded; D004268:DNA-Binding Proteins; D012808:Sigma Factor; D014157:Transcription Factors; C092468:spoIIR protein, Bacillus subtilis; C030069:spore-specific proteins, Bacillus; D003851:Deoxyribonucleases; D004262:DNA Restriction Enzymes