Cloning and nucleotide sequencing of genes for small, acid-soluble spore proteins of Bacillus cereus, Bacillus stearothermophilus, and "Thermoactinomyces thalpophilus"

Loshon, Charles A.; Fliss, E R; Setlow, Barbara; Foerster, Harold F.; Setlow, Peter

doi:10.1128/jb.167.1.168-173.1986

Cited by 37 publications

(48 citation statements)

References 30 publications

(19 reference statements)

Supporting

Mentioning

Contrasting

Order By: Relevance

“…The sources of all of the Escherichia coli strains used, as well as B. subtilis 168, have been described previously (3, 4, * Corresponding author. 9, 14,15). The sources of the pUC plasmids, pJH101 (6) and XgtlO, have also been described previously (3).…”

Section: Methodsmentioning

confidence: 99%

“…B. subtilis chromosomal DNA was isolated and purified as previously described (3). Plasmid or chromosomal DNA was isolated from cells grown overnight at 37°C in 2x YT medium (14) supplemented with appropriate drugs (50 ,ug of ampicillin per ml; 10 pg of tetracycline per ml, 10 ,g of chloramphenicol per ml for E. coli strains, 5 ,ug of chloramphenicol per ml for B. subtilis strains) by using the method of Birnboim and Doly (1); the DNA was purified by CsCl gradient centrifugation if necessary.…”

Section: Methodsmentioning

confidence: 99%

See 1 more Smart Citation

Cloning, nucleotide sequencing, and genetic mapping of the gene for small, acid-soluble spore protein gamma of Bacillus subtilis

Hackett¹,

Setlow²

1987

J Bacteriol

Self Cite

View full text Add to dashboard Cite

The BaciUus subtilis gene (sspE) which codes for small acid-soluble spore protein -y (SASP-,y) was cloned, and its chromosomal location (650, linked to glpD) and nucleotide sequence were determined. The amino acid sequence of SASP--y is similar to that of SASP-B of Bacillus megaterium, but these sequences are not as highly conserved across species as are those of other SASPs. The SASP--y gene is transcribed only in sporulation in parallel with other SASP genes and gives a single mRNA that is -340 nucleotides long. The results of hybridization of an sspE gene probe to Southern blots of B. subtilis DNA suggested that there is only a single gene coding for the SASP-,y type of protein in B. subtilis. This was confirmed by introducing a deletion mutation into the cloned sspE gene and transferring the deletion into the B. subtilis chromosome, with concomitant loss of the wild-type gene. This sspE deletion strain sporulated well, but lacked the SASP-y type of protein.

show abstract

Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Cloning, nucleotide sequencing, and genetic mapping of the gene for small, acid-soluble spore protein gamma of Bacillus subtilis

Hackett¹,

Setlow²

1987

J Bacteriol

Self Cite

View full text Add to dashboard Cite

show abstract

“…Antibiotics were added to the following final concentrations: ampicillin, 50 ,g/ml; chloramphenicol, 3 ,ug/ml for B. subtilis and 5 ,ug/ml for E. coli; and kanamycin, 10 ,ug/ml. Plasmid DNA was isolated from E. coli cells or from B. subtilis cells and spores as previously described (9,19) and purified by CsCl density gradient centrifugation. Plasmid topoisomer distributions were analyzed by electrophoresis in agarose gels containing chloroquine (8 ,ug/ml), and Tav values (average number of negative supertwists) were determined from these gels (19).…”

Section: Methodsmentioning

confidence: 99%

“…1A; Table 3) except for the amino-terminal methionine, which is presumably removed posttranslationally. Since a/P-type SASP show the most sequence heterogeneity in their amino-terminal coding regions (10,25) 9 .Asn (52) 10.Asn (86) 1 .Asn (106) 12 .…”

Section: Constructionmentioning

confidence: 99%

“…The remaining 7.8-kb fragment was ligated with the 0.6-kb HindIII fragment containing the wild-type or mutant sspC gene after the HindlIl ends had been filled in with the large fragment of E. coli DNA polymerase. This mixture was used to transform E. coli to ampicillin resistance; plasmids carrying the sspC gene were identified by colony hybridization with an sspC gene probe (9,17), and plasmids with the sspC gene in the correct orientation with respect to the sspB promoter were identified by restriction enzyme digestion. These plasmids were then cleaved with BamHI, the pUC19 fragment was removed, and the remaining 5.7-kb fragment was ligated and used to transform B. subtilis strains to kanamycin resistance.…”

Section: Tgctggagaaatc-3' and 5'-agaaatcactcaacgmentioning

confidence: 99%

See 1 more Smart Citation

Effects of mutant small, acid-soluble spore proteins from Bacillus subtilis on DNA in vivo and in vitro

1991

Self Cite

View full text Add to dashboard Cite

o$/0-type small, acid-soluble spore proteins (SASP) of Bacillus subtilis bind to DNA and alter its conformation, topology, and photochemistry, and thereby spore resistance to UV light. Three mutations have been introduced into the B. subtilis sspC gene, which codes for the a/$-type wild-type SASP, SspCwt. One mutation (SSpCTYr) was a conservative change, as residue 29 (Leu) was changed to Tyr, an amino acid found at this position in other o/I0-type SASP. The other mutations changed residues conserved in all a/0-type SASP.In one (SspCAIa), residue 52 (Gly) was changed to Ala; in the second (SSpCGIn), residue 57 (Lys) was changed to Gln. The effects of the wild-type and mutant SspC on DNA properties were examined in vivo in B. subtilis spores and Escherichia coli as well as in vitro with use of purified protein. Both SspCwt and SSpCTYr interacted similarly with DNA in vivo and in vitro, restoring much UV resistance to spores lacking major a/l/-type SASP, causing a large increase in plasmid negative supercoiling, and altering DNA UV photochemistry from cell type to spore type. In contrast, SspCAIa had no detectable effect on DNA properties in vivo or in vitro, while SspCGln had effects intermediate between those of SSpCAla and SspCwt. Strikingly, neither SspCAla nor SSpCGln bound well to DNA in vitro. These results confirm the importance of the conserved primary sequence of a/,-type SASP in the ability of these proteins to bind to spore DNA and cause spore UV resistance.Approximately 5 to 12% of the protein of dormant spores of Bacillus and Clostridium species is a group of small, acid-soluble proteins (SASP) of the ot/r type (25). These small proteins (60 to 72 residues) are coded for by a multigene family of at least seven members (termed ssp genes) in Bacillus species. The a/a-type SASP are synthesized in parallel only in the developing forespore during stage III of sporulation and are rapidly degraded during spore germination. While all ssp genes appear to be expressed, generally two (coding for SASP-ot and -1 in Bacillus subtilis) are expressed at high levels, with others expressed at lower levels. One function of a(/a-type SASP is to provide, by their degradation, amino acids for protein synthesis and metabolism early in spore germination. These proteins also have a second role in the resistance of spores to UV light (26). A variety of studies, both in vivo and in vitro, have shown that oa/,-type SASP play a major role in determining the structure of spore DNA, undoubtedly by binding to the DNA and changing its conformation from the B form to A form, with an attendant change in DNA photochemistry which results in spore UV resistance (5,13,(16)(17)(18)(19)23).That a/,8-type SASP are DNA structural proteins is consistent with the extreme conservation of their primary sequence, both within and across species (10,25). Within the Bacillus line of gram-positive spore formers, the sequences of 18 different a/,3-type SASP can be aligned without introduction of gaps; these sequences have 26 identical residues as well ...

show abstract

The p32K Structural Protein of the Atadenovirus Might Have Bacterial Relatives

Élö

Farkas

Dán³

et al. 2003

Journal of Molecular Evolution

View full text Add to dashboard Cite

The primary structure of a novel adenoviral protein referred to as p32K and found exclusively in members of the proposed new genus Atadenovirus was analyzed. The p32K gene sequence was determined from two bovine and one snake adenovirus types. Altogether five different p32K sequences were examined, two of them were obtained from the Gene Bank. The C-terminal part of the protein is conserved and shares similarity with certain bacterial small acid soluble proteins (SASPs). The sequence similarity seems coupled with functional relatedness, i.e. both protein groups are found in structures where the genome of the "dormant" organism is packaged in tight nucleoprotein complexes. In these complexes the DNA is protected against harmful environmental effects until the new reproductive cycle is started with specific protease cleavage of the packaging proteins. Although there is no experimental clue about the role of the p32K proteins, we hypothesize phylogenetic relationship between the two protein groups based on the sequence similarity and the supposed functional similarity. The alignments of these protein groups shows that the conserved part of the p32Ks probably is the result of the duplication of a shorter sequence similar to the SASPs of the Bacilli.

show abstract

Cloning and nucleotide sequencing of genes for small, acid-soluble spore proteins of Bacillus cereus, Bacillus stearothermophilus, and "Thermoactinomyces thalpophilus"

Cited by 37 publications

References 30 publications

Cloning, nucleotide sequencing, and genetic mapping of the gene for small, acid-soluble spore protein gamma of Bacillus subtilis

Cloning, nucleotide sequencing, and genetic mapping of the gene for small, acid-soluble spore protein gamma of Bacillus subtilis

Effects of mutant small, acid-soluble spore proteins from Bacillus subtilis on DNA in vivo and in vitro

The p32K Structural Protein of the Atadenovirus Might Have Bacterial Relatives

Contact Info

Product

Resources

About