YvcPQ is one of the two-component signal transduction systems that respond to specific stimuli and enable cells to adjust multiple cellular functions. It consists of a histidine kinase YvcQ and a response regulator YvcP. In this study, through searching the consensus sequence recognized by YvcP, we found four YvcP-binding motifs in the promoter regions of genes yvcR (BMB171_C4100), BMB171_C4385, kapD (BMB171_C4525) and BMB171_C4835 in Bacillusthuringiensis BMB171 which is a representative of Bacillus cereus group, and confirmed that these genes are regulated by YvcP. We compared the sequence of yvcPQ and its downstream genes in genus Bacillus, and found two different kinds of yvc locus, one was the yvcPQ-RS in B. subtilis species and the other was the yvcPQ-R-S1S2 in B. cereus group. Furthermore, we found that YvcP activates the transcription of yvcS1S2 (downstream of yvcR) to promote bacterial resistance to bacitracin and deletion of either yvcPQ operon or yvcS1S2 operon renders the bacterial cells more sensitive to bacitracin. This study enriched our understanding of both the YvcPQ’s function and the mechanism of bacterial resistance to bacitracin.

YvqEC is one of the two-component signal transduction systems that may respond to cell envelope stress and enable cells to adjust multiple cellular functions. It consists of a histidine kinase YvqE and a response regulator YvqC. In this study, we separately constructed a single gene mutant ΔyvqE and a double gene mutant ΔyvqEC in Bacillusthuringiensis BMB171 through a homing endonucleases I-SceI mediated markerless gene deletion method. We found that the deletion of either yvqE or yvqEC weakened the resistance of B.thuringiensis against vancomycin. We also identified nine operons that may be involved in the cellular metabolism regulated by YvqC. This study not only enriches our understanding of bacterial resistance mechanisms against vancomycin, but also helps investigate the functions of YvqEC.

Bacillus thuringiensis, a Gram-positive endospore-forming bacterium, is characterized by the formation of parasporal crystals consisting of insecticidal crystal proteins (ICPs) during sporulation. We reveal gene expression profiles and regulatory mechanisms associated with spore and parasporal crystal formation based on transcriptomics and proteomics data of B. thuringiensis strain CT-43. During sporulation, five ICP genes encoded by CT-43 were specifically transcribed; moreover, most of the spore structure-, assembly-, and maturation-associated genes were specifically expressed or significantly up-regulated, with significant characteristics of temporal regulation. These findings suggest that it is essential for the cell to maintain efficient operation of transcriptional and translational machinery during sporulation. Our results indicate that the RNA polymerase complex δ and ω subunits, cold shock proteins, sigma factors, and transcriptional factors as well as the E2 subunit of the pyruvate dehydrogenase complex could cooperatively participate in transcriptional regulation via different mechanisms. In particular, differences in processing and modification of ribosomal proteins, rRNA, and tRNA combined with derepression of translational inhibition could boost the rate of ribosome recycling and assembly as well as translation initiation, elongation, and termination efficiency, thereby compensating for the reduction in ribosomal levels. The efficient operation of translational machineries and powerful protein-quality controlling systems would thus ensure biosyntheses of a large quantity of proteins with normal biological functions during sporulation.

A novel, simple and efficient dispersive liquid–liquid microextraction based on solidification of floating organic droplet (DLLME-SFO) technique coupled with high-performance liquid chromatography with ultraviolet detection (HPLC–UV) and liquid chromatography–tandem mass spectrometry (LC–MS/MS) was developed for the determination of triclosan and its degradation product 2,4-dichlorophenol in real water samples. The extraction solvent used in this work is of low density, low volatility, low toxicity and proper melting point around room temperature. The extractant droplets can be collected easily by solidifying it at a lower temperature. Parameters that affect the extraction efficiency, including type and volume of extraction solvent and dispersive solvent, salt effect, pH and extraction time, were investigated and optimized in a 5 mL sample system by HPLC–UV. Under the optimum conditions (extraction solvent: 12 μL of 1-dodecanol; dispersive solvent: 300 of μL acetonitrile; sample pH: 6.0; extraction time: 1 min), the limits of detection (LODs) of the pretreatment method combined with LC–MS/MS were in the range of 0.002–0.02 μg L−1 which are lower than or comparable with other reported approaches applied to the determination of the same compounds. Wide linearities, good precisions and satisfactory relative recoveries were also obtained. The proposed technique was successfully applied to determine triclosan and 2,4-dichlorophenol in real water samples.

Although the tetracysteine (TC) motif has been used as a tag, the binding stability between TC motif and biarsenical reagent against extreme conditions as well as its capacity as a quantitative tag remains not well developed. To reveal these problems, we chose enoyl−acyl carrier protein reductase (FabI), which was involved in the final step of elongation in the bacterial fatty acid biosynthesis, to be tagged by the TC motif. Taking enhanced green fluorescent protein (EGFP) tagged FabI as a control, we investigated the activities of various TC tagged FabIs (N-terminus, C-terminus, or both N- and C-terminus TC motif). The results showed that all the TC tagged FabIs had high enzyme activities while the EGFP tagged FabI exhaustively lost the activity. Beside this, the characteristics of the tag, including labeling stability against extreme conditions, capacity for quantitative analysis, and ability for in-cell labeling, were also investigated. We demonstrated for the first time that the binding between FlAsH reagent and TC motif was stable against high pressure, high field strength, high temperature, and ultrasound. Furthermore, we verified the potential of TC motif for quantitative analysis of target protein by different approaches, including SDS−PAGE, spectrofluorometry (SPF), and capillary zone electrophoresis (CZE).

By structure-based virtual screening and experimental verification, two Chinese medicine monomers, luteolin and curcumin, had been proved to be uncompetitive inhibitors of enoyl-ACP reductase from Escherichia coli (EcFabI) with the inhibition constant (Ki) of 7.1 μM and 15.0 μM, respectively. In particular, curcumin had apparent antibacterial activity against E. coli, and the minimum inhibition concentration (MIC90) was 73.7 μg/mL. Importantly, fabI-overexpressing E. coli showed reduced susceptibility to the inhibitor compared with the wild-type strains, demonstrating that its antibacterial action is mediated by the inhibition of EcFabI.Curcumin has been proved to be an uncompetitive inhibitor of enoyl-ACP reductase from Escherichia coli (EcFabI) with the inhibition constant (Ki) of 15 μM. Particularly, curcumin had apparent antibacterial activity against E. coli, and the corresponding MIC90 was 73.7 μg/mL.