The Single‐stranded‐DNA‐binding Proteins (SSB) of Proteus mirabilis and Serratia marcescens

Abstract

The single-stranded-DNA-binding (SSB) proteins from Proteus mirabilis and Serratia marcescens were purified from overproducing Escherichia coli strains, which were devoid of their own ssb gene. The strains harboured an endA insertion mutation and a xonA mutation resulting in the absence of endonuclease I and exonuclease I activities from the preparations. The amino acid sequences of the SSB of all three species are nearly identical in the N-terminal parts of the proteins that contain the DNA-binding domain, but differ in the C-terminal parts. Both proteins have an apparent binding-site size of 65 and 35 nucleotides at high and low salt concentrations, respectively. The association-rate constant for binding to poly(dT) is 3.2 x 10(8) M-1 s-1 for P. mirabilis SSB (PmiSSB) and 3.4 x 10(8) M-1 s-1 for S. marcescens SSB (SmaSSB). These binding parameters are very similar to those of E. coli SSB (EcoSSB). The structural similarity of the proteins is also documented by the finding that they can exchange subunits among each other to form mixed tetramers. The transcriptional regulation of the ssb and uvrA genes from P. mirabilis and S. marcescens in SOS-induced E. coli cells was studied using lacZ fusions. While the uvrA genes were inducible, there was no induction of the ssb genes transcribed divergently from the uvrA genes. Apparently, regions with nucleotide sequence similarity to the E. coli SOS-box preceding the ssb genes of P. mirabilis and S. marcescens had no gross effect on the transcription. Studies on growth of the cells and recovery from ultraviolet damage indicate that the heterologous SSB proteins support DNA replication and recombinational DNA repair of E. coli with the same efficiency as the E. coli SSB protein. Interactions with other E. coli proteins involved in these processes either do not occur, or are not impeded.

Differential Expression of Two Paralogous Genes of Bacillus subtilis Encoding Single-Stranded DNA Binding Protein

“…The same organization was found for the uvrA and ssb genes in Sinorhizobium meliloti (24). Although this gene organization is also identical in Proteus mirabilis and Serratia marcescens, the ssb genes of these bacteria are not inducible by DNA damage (7). It has been suggested that E. coli SSB negatively autoregulates its own translation, because it is capable of binding to its own mRNA, in this way inhibiting translation (21).…”

“…Notably, YwpH is lacking 66 amino acid residues of the C terminus of SSB. Although the amino acid sequences of bacterial SSBs are highly conserved within the first two thirds of the protein containing the DNA-binding domains, they diverge substantially in the C-terminal third region (7). The C-terminal region of E. coli SSB is not required for DNA binding in vitro, but is essential for its in vivo function (6,29).…”

The Euryarchaeota, Nature's Medium for Engineering of Single-stranded DNA-binding Proteins

“…In ssDNA-binding proteins, the OB folds typically comprise ϳ100 amino acids (21), and although the majority of bacterial SSB genes code for products that contain a single OB fold, their active forms are homotetramers (22)(23)(24). Most interestingly, in Deinococcus radiodurans (25) and Thermus aquaticus (26), genes encoding SSBs with two OB folds have been described recently.…”

Crystal Structure of PriB, a Primosomal DNA Replication Protein of Escherichia coli

“…Interestingly, bacterial SSBs from different species can form cross-species heterotetramers through their six-stranded ␤-sheet surfaces (43). This finding suggests that the similar six-stranded ␤-sheet surfaces of the PriB dimer could interact with an EcoSSB dimer.…”