N-terminal Amino Acid Residues Mediate Protein-Protein Interactions between DNA-bound α/β-Type Small, Acid-soluble Spore Proteins from Bacillus Species

Self Cite

A derivative of SspC, a minor ␣/␤-type, small, acid-soluble spore protein (SASP) from Bacillus subtilis, was generated that has a very high affinity for DNA. This protein (SspC ⌬11-D13K ) was able to confer UV resistance on spores lacking ␣/␤-type SASP, and spores with SspC ⌬11-D13K triggered germination normally. However, SspC ⌬11-D13K blocked outgrowth of >90% of germinated spores, and SspC ⌬11-D13K persisted in these germinated spores, whereas wild-type SspC was almost completely degraded. The outgrowth phenotype of spores with SspC ⌬11-D13K is proposed to be due to the high stability of the SspC ⌬11-D13K -DNA complex, which prevents rapid degradation of this ␣/␤-type SASP early in germination. The persistence of this protein on spore DNA then interferes with transcription during spore outgrowth.

Section: ⌬11-d13kmentioning

confidence: 99%

An α/β-Type, Small, Acid-Soluble Spore Protein Which Has Very High Affinity for DNA Prevents Outgrowth of Bacillus subtilis Spores

Hayes

Setlow

2001

Self Cite

“…The stoichiometry for the binding of this protein to pdG:pdC was 1 protein per 4 bp, similar to what was found for SspC wt (8,16). Because of the very tight binding of SspC ⌬N11-D13K-C3 and SspC ⌬N11-D13K to pUC19, the DNA used routinely to quantitate protein-DNA binding was pdAdT: pdAdT, a polynucleotide for which ␣/␤-type SASP have lower affinity than for pUC19 (4,6). At pH 7, SspC…”

Section: Effects Of Deletion Of C-terminal Residues From Sspcmentioning

confidence: 91%

“…SspC ⌬C9 , in which five residues beyond glycine 68 are deleted, is not accumulated in E. coli. We presume, as found for SspC variants with very large N-terminal deletions (4), that removal of too much of the C-terminal region of SspC wt , as in SspC ⌬C9 , results in a protein that is rapidly degraded, since the protein cannot bind DNA and is thus not protected against proteolysis (27). C-terminal extensions of SspC wt , however, appear to increase DNA binding.…”

Section: Discussionmentioning

confidence: 99%

“…Previous studies focusing on the N-terminal region of SspC wt identified some key residues affecting DNA binding (11,33). The culmination of these studies was the production of the SspC ⌬N11-D13K variant, in which 11 residues at the N terminus were deleted while the positive charge in this region of the protein was retained; these changes resulted in a greater-than-10-fold improvement in the protein's affinity for pUC19 DNA over that of SspC wt (4,8). The SspC ⌬N11-D13K variant also provided far greater protection to DNA against deamination and restriction enzyme digestion in vitro than did SspC wt (34).…”

mentioning

confidence: 99%

See 1 more Smart Citation

Effects of Carboxy-Terminal Modifications and pH on Binding of a Bacillus subtilis Small, Acid-Soluble Spore Protein to DNA

Kosman

Setlow

2003

Self Cite

Variants of the wild-type Bacillus subtilis ␣/␤-type small, acid-soluble spore protein (SASP) SspC wt were designed to evaluate the contribution of C-terminal residues to these proteins' affinity for DNA. SspC variants lacking one to three C-terminal residues were similar to SspC wt in DNA binding, but removal of six C-terminal residues greatly decreased DNA binding. In contrast, a C-terminal extension of three residues increased SspC's affinity for DNA 5-to 10-fold. C-terminal and N-terminal changes that independently caused large increases in SspC-DNA binding affinity were combined and produced an additive effect on DNA binding; the affinity of the resulting variant, SspC ⌬N11-D13K-C3 , for DNA was increased >20-fold over that of SspC wt . For most of the SspC variants tested, lowering the pH from 7 to 6 improved DNA binding two-to sixfold, although the opposite effect was observed with variants having additional C-terminal basic residues. In vitro, the binding of SspC ⌬N11-D13K-C3 to DNA suppressed the formation of cyclobutane-type thymine dimers and promoted the formation of the spore photoproduct upon UV irradiation to the same degree as the binding of SspC wt . However, B. subtilis spores lacking major ␣/␤-type SASP and overexpressing SspC ⌬N11-D13K-C3 had a 10-foldlower viability and far less UV and heat resistance than spores overexpressing SspC wt . This apparent lack of DNA protection by SspC ⌬N11-D13K-C3 in vivo is likely due to the twofold-lower level of this protein in spores compared to the level of SspC wt , perhaps because of effects of SspC ⌬N11-D13K-C3 on gene expression in the forespore during sporulation. The latter results indicate that only moderately strong binding of ␣/␤-type SASP to DNA is important to balance the potentially conflicting requirements for these proteins in DNA transcription and DNA protection during spore formation, spore dormancy, and spore germination and outgrowth.

“…SASPs can comprise 5 to 20% of the total spore protein in B. subtilis and are capable of saturation binding of spore DNA (28). These proteins are devoid of secondary structures, which allows for solubility at low pH (11). However, SASPs become significantly ␣-helical upon binding to DNA (38,45).…”

mentioning

confidence: 99%

Small Acid-Soluble Proteins with Intrinsic Disorder Are Required for UV Resistance in Myxococcus xanthus Spores

Dahl

Fordice

2011

Bacterial sporulation in Gram-positive bacteria results in small acid-soluble proteins called SASPs that bind to DNA and prevent the damaging effects of UV radiation. Orthologs of Bacillus subtilis genes encoding SASPs can be found in many sporulating and nonsporulating bacteria, but they are noticeably absent from sporeforming, Gram-negative Myxococcus xanthus. This is despite the fact that M. xanthus can form UV-resistant spores. Here we report evidence that M. xanthus produces its own unique group of low-molecular-weight, acid-soluble proteins that facilitate UV resistance in spores. These M. xanthus-specific SASPs vary depending upon whether spore formation is induced by starvation inside cell aggregations of fruiting bodies or is induced artificially by glycerol induction. Molecular predictions indicate that M. xanthus SASPs may have some association with the cell walls of M. xanthus spores, which may signify a different mechanism of UV protection than that seen in Gram-positive spores.