The proteome of growing cells of Bacillus subtilis was analyzed in order to provide the basis for its application in microbial physiology. DNA arrays were used to calculate the number of genes transcribed in growing cells. From the 4100 B. subtilis genes, 2515 were actively transcribed in cells grown under standard conditions. From these genes 1544 proteins should be covered by our standard gel system pI 4-7. Using this standard gel system and supplementary zoom gels (pI 5.5-6.7, 5-6, 4.5-5.5, and 4-5) 693 proteins which are expressed in growing cells were detected that cover more than 40% of the vegetative proteome predicted for this region. Particularly broad coverage and thus comprehensive monitoring will be possible for central carbohydrate metabolism (glycolysis, pentose phosphate shunt, and citric acid cycle), amino acid synthesis pathways, purine and pyrimidine metabolism, fatty acid metabolism, and main cellular functions like replication, transcription, translation, and cell wall synthesis. Comparing the theoretical pI and Mr values with those experimentally determined a reasonable correlation was found for the majority of protein spots. By a color code outliers with dramatic deviations in charge or mass were visualized that may indicate post-translational modifications. In addition to the cytosolic neutral and alkaline proteins, 130 membrane proteins were found relying on sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) separation in combination with electrospray ionization-tandem mass spectrometry (ESI-MS/MS) techniques. The vegetative proteome containing 876 proteins in total is now ready for physiological applications. Two main proteome fractions (pI 4-7 and zoom gel pI 4.5-5.5) should be sufficient for such high-throughput physiological proteomics.

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Aromatic organic compounds that are present in the environment can have toxic effects or provide carbon sources for bacteria. We report here the global response of Bacillus subtilis 168 to phenol and catechol using proteome and transcriptome analyses. Phenol induced the HrcA, sigmaB and CtsR heat-shock regulons as well as the Spx disulfide stress regulon. Catechol caused the activation of the HrcA and CtsR heat-shock regulons and a thiol-specific oxidative stress response involving the Spx, PerR and FurR regulons but no induction of the sigmaB regulon. The most surprising result was that several catabolite-controlled genes are derepressed by catechol, even if glucose is taken up under these conditions. This derepression of the carbon catabolite control was dependent on the glucose concentration in the medium, as glucose excess increased the derepression of the CcpA-dependent lichenin utilization licBCAH operon and the ribose metabolism rbsRKDACB operon by catechol. Growth and viability experiments with catechol as sole carbon source suggested that B. subtilis is not able to utilize catechol as a carbon-energy source. In addition, the microarray results revealed the very strong induction of the yfiDE operon by catechol of which the yfiE gene shares similarities to glyoxalases/bleomycin resistance proteins/extradiol dioxygenases. Using recombinant His6-YfiE(Bs) we demonstrate that YfiE shows catechol-2,3-dioxygenase activity in the presence of catechol as the metabolite 2-hydroxymuconic semialdehyde was measured. Furthermore, both genes of the yfiDE operon are essential for the growth and viability of B. subtilis in the presence of catechol. Thus, our studies revealed that the catechol-2,3-dioxygenase YfiE is the key enzyme of a meta cleavage pathway in B. subtilis involved in the catabolism of catechol.

In this paper we have defined proteome signatures of Bacillus subtilis in response to heat, salt, peroxide, and superoxide stress as well as after starvation for ammonium, tryptophan, glucose, and phosphate using the 2-D gel-based approach. In total, 79 stress-induced and 155 starvation-induced marker proteins were identified including 50% that are not expressed in the vegetative proteome. Fused proteome maps and a color coding approach have been used to define stress-specific regulons that are involved in specific adaptative functions (HrcA for heat, PerR and Fur for oxidative stress, RecA for peroxide, CymR and S-box for superoxide stress). In addition, starvation-specific regulons are defined that are involved in the uptake or utilization of alternative nutrient sources (TnrA, sigmaL/BkdR for ammonium; tryptophan-activated RNA-binding attenuation protein for tryptophan; CcpA, CcpN, sigmaL/AcoR for glucose; PhoPR for phosphate starvation). The general stress or starvation proteome signatures include the CtsR, Spx, sigmaL/RocR, sigmaB, sigmaH, CodY, sigmaF, and sigmaE regulons. Among these, the Spx-dependent oxidase NfrA was induced by all stress conditions indicating stress-induced protein damages. Finally, a subset of sigmaH-dependent proteins (sporulation response regulator, YvyD, YtxH, YisK, YuxI, YpiB) and the CodY-dependent aspartyl phosphatase RapA were defined as general starvation proteins that indicate the transition to stationary phase caused by starvation.

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OA and OLA coated CoFe2O4 nanoparticles encapsulated with PMAO through hydrophobic interactions were developed for hyperthermia applications.

Phenolic acids that are present in plant-soil ecosystems can be considered as toxins which induce specific stress responses in microorganisms. In this paper, we have analyzed the global response of the soil bacterium Bacillus subtilis to salicylic acid using proteomics and transcriptomics. The results demonstrate that salicylic acid caused predominantly the induction of the SigmaB-dependent general stress response in B. subtilis which is not related to the acidic conditions. Treatment of B. subtilis with growth-inhibitory concentrations of 4 mM salicylic acid caused protein damage in B. subtilis as reflected by the induction of the CtsR and Spx regulons. Both phenolic acid decarboxylases (pads) of B. subtilis padC and bsdBCD (yclBCD) were induced by 4 mM salicylic acid that were previously shown to be involved in decarboxylation and detoxification of different phenolic acids. Deletion of the putative LysR-type regulator encoded by the divergently transcribed bsdA (yclA) gene upstream of the bsdBCD operon revealed that BsdA is the transcriptional activator of bsdBCD expression in response to salicylic acid. Phenotype analysis of bsdA and padC single and double mutants demonstrated that both pads confer resistance to salicylic acid in B. subtilis.

Quercus gilva Bume (QGB, family Fagaceae) is a tall evergreen oak species tree that grows in warm temperate regions in Korea, Japan, China and Taiwan. Quercus plants have long been the basis of traditional medicines. Their clinical benefits according to traditional medicine include relief of urolithiasis, tremors and inflammation. In the present study, the anti-urolithiasis activity including anti-inflammatory and anti-oxidative activities, of some phenolic compounds isolated from QGB were described. Seven compounds were isolated and identified as picraquassioside D (1), quercussioside (2), (+)-lyoniresinol-9′α-O-β-d-xylopyranoside (3), (+)-catechin (4), (−)-epicatechin (5), procyanidin B-3 (6), and procyanidin B-4 (7). Compounds 5–7 showed potent anti-oxidative and anti-inflammatory activities. These compounds were further tested for their inhibition of the gene expression of the inflammatory cytokines. The three compounds 5–7 showed dose-dependent inhibitory activities on gene expression of COX-2 and IL-1β. In vivo, urolithiasis was induced more effectively in an animal model of acute urolithiasis by the administration of QGB extract. These results indicate the potential of compounds from QGB in the treatment of urolithiasis.