Biomedical subjects

Sierd Bron

Publications and source records attributed to Sierd Bron.

25 records · Page 2Linked to original sources

PubMed2002

Thiol-disulfide oxidoreductases are essential for the production of the lantibiotic sublancin 168.

Thiol-disulfide oxidoreductases are required for disulfide bond formation in proteins that are exported from the cytoplasm. Four enzymes of this type, termed BdbA, BdbB, BdbC, and BdbD, have been identified in the Gram-positive eubacterium Bacillus subtilis. BdbC and BdbD have been shown to be critical for the folding of a protein required for DNA uptake during natural competence. In contrast, no function has been assigned so far to the BdbA and BdbB proteins. The bdbA and bdbB genes are located in one operon that also contains the genes specifying the lantibiotic sublancin 168 and the ATP-binding cassette transporter SunT. Interestingly sublancin 168 contains two disulfide bonds. The present studies demonstrate that SunT and BdbB, but not BdbA, are required for the production of active sublancin 168. In addition, the BdbB paralogue BdbC is at least partly able to replace BdbB in sublancin 168 production. These observations show the unprecedented involvement of thiol-disulfide oxidoreductases in the synthesis of a peptide antibiotic. Notably BdbB cannot complement BdbC in competence development, showing that these two closely related thiol-disulfide oxidoreductases have different, but partly overlapping, substrate specificities.

ATP-Binding Cassette Transporters↗

PubMed2002

A novel class of heat and secretion stress-responsive genes is controlled by the autoregulated CssRS two-component system of Bacillus subtilis.

Bacteria need dedicated systems that allow appropriate adaptation to the perpetual changes in their environments. In Bacillus subtilis, two HtrA-like proteases, HtrA and HtrB, play critical roles in the cellular response to secretion and heat stresses. Transcription of these genes is induced by the high-level production of a secreted protein or by a temperature upshift. The CssR-CssS two-component regulatory system plays an essential role in this transcriptional activation. Transcription of the cssRS operon is autoregulated and can be induced by secretion stress, by the absence of either HtrA or HtrB, and by heat stress in a HtrA null mutant strain. Two start sites are used for cssRS transcription, only one of which is responsive to heat and secretion stress. The divergently transcribed htrB and cssRS genes share a regulatory region through which their secretion and heat stress-induced expression is linked. This study shows that CssRS-regulated genes represent a novel class of heat-inducible genes, which is referred to as class V and currently includes two genes: htrA and htrB.

Bacillus subtilis↗

PubMed2001

The bdbDC operon of Bacillus subtilis encodes thiol-disulfide oxidoreductases required for competence development.

The development of genetic competence in the Gram-positive eubacterium Bacillus subtilis is a complex postexponential process. Here we describe a new bicistronic operon, bdbDC, required for competence development, which was identified by the B. subtilis Systematic Gene Function Analysis program. Inactivation of either the bdbC or bdbD genes of this operon results in the loss of transformability without affecting recombination or the synthesis of ComK, the competence transcription factor. BdbC and BdbD are orthologs of enzymes known to be involved in extracytoplasmic disulfide bond formation. Consistent with this, BdbC and BdbD are needed for the secretion of the Escherichia coli disulfide bond-containing alkaline phosphatase, PhoA, by B. subtilis. Similarly, the amount of the disulfide bond-containing competence protein ComGC is severely reduced in bdbC or bdbD mutants. In contrast, the amounts of the competence proteins ComGA and ComEA remain unaffected by bdbDC mutations. Taken together, these observations imply that in the absence of either BdbC or BdbD, ComGC is unstable and that BdbC and BdbD catalyze the formation of disulfide bonds that are essential for the DNA binding and uptake machinery.

Alkaline Phosphatase↗

PubMed2001

Detergent-independent in vitro activity of a truncated Bacillus signal peptidase.

The Gram-positive eubacterium Bacillus subtilis contains five chromosomally encoded type I signal peptidases (SPases) for the processing of secretory pre-proteins. Even though four of these SPases, denoted SipS, SipT, SipU and SipV, are homologous to the unique SPase I of Escherichia coli, they are structurally different from that enzyme, being almost half the size and containing one membrane anchor instead of two. To investigate whether the unique membrane anchor of Bacillus SPases is required for in vitro activity, soluble forms of SipS of B. subtilis, SipS of Bacillus amyloliquefaciens and SipC of the thermophile Bacillus caldolyticus were constructed. Of these three proteins, only a hexa-histidine-tagged soluble form of SipS of B. amyloliquefaciens could be isolated in significant quantities. This protein displayed optimal activity at pH 10, which is remarkable considering the fact that the catalytic domain of SPases is located in an acidic environment at the outer surface of the membrane of living cells. Strikingly, in contrast to what has been previously reported for the soluble form of the E. coli SPase, soluble SipS was active in the absence of added detergents. This observation can be explained by the fact that a highly hydrophobic surface domain of the E. coli SPase, implicated in detergent-binding, is absent from SipS.

Amino Acid Sequence↗

PubMed2000

Chaperone-like activities of the CsaA protein of Bacillus subtilis.

The growth and protein export defects of Escherichia coli secA51(Ts) strains can be suppressed by the CsaA protein of Bacillus subtilis. The present studies indicate that this effect can be attributed to chaperone-like activities of CsaA. First, CsaA stimulated protein export in secB, groES and dnaJ mutant strains of E. coli. Second, CsaA suppressed the growth defects of dnaK, dnaJ and grpE mutants of E. coli. Third, and most importantly, CsaA exhibited chaperone-like properties by stimulating the reactivation of heat-denatured firefly luciferase in groEL, groES, dnaK and grpE mutant strains of E. coli, and by preventing the aggregation of heat-denatured luciferase in vitro. Thus, it seems that CsaA suppresses the growth and secretion defects of E. coli secA(Ts) strains either by improving the translocation competence of exported pre-proteins, thereby making them better substrates for mutant SecA proteins, or by stimulating the translocation activity of mutant SecA proteins.

Adenosine Triphosphatases↗

PubMed1998

The 172 kb prkA-addAB region from 83 degrees to 97 degrees of the Bacillus subtilis chromosome contains several dysfunctional genes, the glyB marker, many genes encoding transporter proteins, and the ubiquitous hit gene.

A 171812 bp nucleotide sequence between prkA and addAB (83 degrees to 97 degrees) on the genetic map of the Bacillus subtilis 168 chromosome was determined and analysed. An accurate physical/genetic map of this previously poorly described chromosomal region was constructed. One hundred and seventy open reading frames (ORFs) were identified on the DNA fragment. These include the previously described genes cspB, glpPFKD, spoVR, phoAIV, papQ, citRA, sspB, prsA, hpr, pbpF, hemEHY, aprE, comK and addAB. ORF yhaF in this region corresponds to the glyB marker. Among the striking features of this region are: an abundance of genes encoding (putative) transporter proteins, several dysfunctional genes, the ubiquitous hit gene, and five multidrug-resistance-like genes. These analyses have also revealed the existence of numerous paralogues of ORFs in this region: about two-thirds of the putative genes seem to have at least one paralogue in the B. subtilis genome.

Bacillus subtilis↗

PubMed1997

The signal peptidase II (Isp) gene of Bacillus subtilis.

The gene encoding the type II signal peptidase (SPase II) of Bacillus subtilis was isolated by screening a genomic DNA library of this bacterium for the ability to increase the levels of globomycin resistance in Escherichia coli, and to complement the growth deficiency at the non-permissive temperature of E. coli strain Y815 carrying a temperature-sensitive mutation in its Isp gene for SPase II. The deduced amino acid sequence of the B. subtilis SPase II showed significant similarity with those of other known SPase II enzymes. Activity of the B. subtilis SPase II was demonstrated by a pulse-labelling experiment in E. coli. In B. subtilis, the Isp gene is flanked by the isoleucyl-tRNA synthetase (ileS) gene and the pyrimidine biosynthetic (pyr) gene cluster, which is known to map at 139 degrees of the chromosome. In the Gram-positive bacteria studied thus far, Isp appears to be the first gene in an operon. The promoter-distal gene ("orf4') of this operon specifies a hypothetical protein in bacteria and yeast.

Amino Acid Sequence↗