Enhancement of surfactin production of Bacillus subtilis fmbR by replacement of the native promoter with the Pspac promoter

2009 ◽  
Vol 55 (8) ◽  
pp. 1003-1006 ◽  
Author(s):  
Huigang Sun ◽  
Xiaomei Bie ◽  
Fengxia Lu ◽  
Yaping Lu ◽  
Yundailai Wu ◽  
...  
Keyword(s):  
Bacillus Subtilis ◽  
Bacillus Cereus ◽  
Type Strain ◽  
Wild Type Strain ◽  
Inducible Promoter ◽  
Type B ◽  
Wild Type ◽  
Native Promoter ◽  
Surfactin Production

Bacillus subtilis fmbR-1 was obtained from wild-type B. subtilis fmbR by replacement of the native promoter of the surfactin operon with the inducible promoter Pspac. The recombinant B. subtilis fmbR-1 produced more antibacterial substances than the wild-type strain. The overproducing phenotype was related to the enhancement of antagonistic activities against Bacillus cereus . HPLC peaks of surfactin for recombinant fmbR-1 showed patterns of lipopeptides similar to those of the wild-type strain, and surfactin production of the recombinant fmbR-1 was up to about fivefold greater than that of the wild-type strain without induction by isopropyl β-d-1-thiogalactopyranoside. However, the production of surfactin increased to about 10-fold more than that of the wild-type strain when it was induced by isopropyl β-d-1-thiogalactopyranoside.

scholarly journals Mycosubtilin Overproduction by Bacillus subtilis BBG100 Enhances the Organism's Antagonistic and Biocontrol Activities

2005 ◽  
Vol 71 (8) ◽  
pp. 4577-4584 ◽  
Author(s):  
Valérie Leclère ◽  
Max Béchet ◽  
Akram Adam ◽  
Jean-Sébastien Guez ◽  
Bernard Wathelet ◽  
...  
Keyword(s):  
Bacillus Subtilis ◽  
Type Strain ◽  
Wild Type Strain ◽  
Germination Rate ◽  
Pathogenic Fungi ◽  
Strain Atcc ◽  
Wild Type ◽  
Native Promoter ◽  
Replication Gene ◽  
Biocontrol Potential

ABSTRACT A Bacillus subtilis derivative was obtained from strain ATCC 6633 by replacement of the native promoter of the mycosubtilin operon by a constitutive promoter originating from the replication gene repU of the Staphylococcus aureus plasmid pUB110. The recombinant strain, designated BBG100, produced up to 15-fold more mycosubtilin than the wild type produced. The overproducing phenotype was related to enhancement of the antagonistic activities against several yeasts and pathogenic fungi. Hemolytic activities were also clearly increased in the modified strain. Mass spectrometry analyses of enriched mycosubtilin extracts showed similar patterns of lipopeptides for BBG100 and the wild type. Interestingly, these analyses also revealed a new form of mycosubtilin which was more easily detected in the BBG100 sample. When tested for its biocontrol potential, wild-type strain ATCC 6633 was almost ineffective for reducing a Pythium infection of tomato seedlings. However, treatment of seeds with the BBG100 overproducing strain resulted in a marked increase in the germination rate of seeds. This protective effect afforded by mycosubtilin overproduction was also visualized by the significantly greater fresh weight of emerging seedlings treated with BBG100 compared to controls or seedlings inoculated with the wild-type strain.

NEW LOCI IN DICTYOSTELIUM DISCOIDEUM DETERMINING PIGMENT FORMATION AND GROWTH ON BACILLUS SUBTILIS

Genetics ◽  
1980 ◽  
Vol 96 (1) ◽  
pp. 115-123
Author(s):  
James H Morrissey ◽  
Steven Wheeler ◽  
William F Loomis
Keyword(s):  
Bacillus Subtilis ◽  
Dictyostelium Discoideum ◽  
Type Strain ◽  
Wild Type Strain ◽  
Wild Type ◽  
Linkage Groups ◽  
New Mutation ◽  
Pigment Formation ◽  
Complementation Groups ◽  
First Time

ABSTRACT Seventeen independently isolated pigmentless (white) mutations in Dictyostelium discoideum are all recessive and fall into three complementation groups identifying two new whi loci in addition to the previously characterized whiA locus. whiB and whiC map to linkage groups III and IV, respectively. In addition, it was discovered that our laboratory stock of NC4, the wild-type strain from which these mutants were derived, has spontaneously lost the ability to grow on Bacillus subtilis. This new mutation, bsgB500, maps to linkage group VII and is not allelic to bsgA. bsgB500 is the first spontaneously derived mutation in D. discoideum that can be used to select heterozygous diploids, and for the first time allows genetic analysis to be routinely performed on strains derived from an unmutagenized background.

scholarly journals Selective Pressure for Biofilm Formation in Bacillus subtilis: Differential Effect of Mutations in the Master Regulator SinR on Bistability

mBio ◽  
2018 ◽  
Vol 9 (5) ◽  
Author(s):  
Jan Kampf ◽  
Jan Gerwig ◽  
Kerstin Kruse ◽  
Robert Cleverley ◽  
Miriam Dormeyer ◽  
...  
Keyword(s):  
Gene Expression ◽  
Bacillus Subtilis ◽  
Biofilm Formation ◽  
Type Strain ◽  
Wild Type Strain ◽  
Selective Pressure ◽  
Wild Type ◽  
Master Regulator ◽  
Form Complex ◽  
Content Type

ABSTRACT Biofilm formation by Bacillus subtilis requires the expression of genes encoding enzymes for extracellular polysaccharide synthesis and for an amyloid-like protein. The master regulator SinR represses all the corresponding genes, and repression of these key biofilm genes is lifted when SinR interacts with its cognate antagonist proteins. The YmdB phosphodiesterase is a recently discovered factor that is involved in the control of SinR activity: cells lacking YmdB exhibit hyperactive SinR and are unable to relieve the repression of the biofilm genes. In this study, we have examined the dynamics of gene expression patterns in wild-type and ymdB mutant cells by microfluidic analysis coupled to time-lapse microscopy. Our results confirm the bistable expression pattern for motility and biofilm genes in the wild-type strain and the loss of biofilm gene expression in the mutant. Moreover, we demonstrated dynamic behavior in subpopulations of the wild-type strain that is characterized by switches in sets of the expressed genes. In order to gain further insights into the role of YmdB, we isolated a set of spontaneous suppressor mutants derived from ymdB mutants that had regained the ability to form complex colonies and biofilms. Interestingly, all of the mutations affected SinR. In some mutants, large genomic regions encompassing sinR were deleted, whereas others had alleles encoding SinR variants. Functional and biochemical studies with these SinR variants revealed how these proteins allowed biofilm gene expression in the ymdB mutant strains. IMPORTANCE Many bacteria are able to choose between two mutually exclusive lifestyles: biofilm formation and motility. In the model bacterium Bacillus subtilis, this choice is made by each individual cell rather than at the population level. The transcriptional repressor SinR is the master regulator in this decision-making process. The regulation of SinR activity involves complex control of its own expression and of its interaction with antagonist proteins. We show that the YmdB phosphodiesterase is required to allow the expression of SinR-repressed genes in a subpopulation of cells and that such subpopulations can switch between different SinR activity states. Suppressor analyses revealed that ymdB mutants readily acquire mutations affecting SinR, thus restoring biofilm formation. These findings suggest that B. subtilis cells experience selective pressure to form the extracellular matrix that is characteristic of biofilms and that YmdB is required for the homeostasis of SinR and/or its antagonists.

scholarly journals A Mutation in the 3-Phosphoglycerate Kinase Gene Allows Anaerobic Growth of Bacillus subtilis in the Absence of ResE Kinase

1999 ◽  
Vol 181 (22) ◽  
pp. 7087-7097 ◽  
Author(s):  
Michiko M. Nakano ◽  
Yi Zhu ◽  
Koki Haga ◽  
Hirofumi Yoshikawa ◽  
Abraham L. Sonenshein ◽  
...  
Keyword(s):  
Bacillus Subtilis ◽  
Type Strain ◽  
Wild Type Strain ◽  
Response Regulator ◽  
Anaerobic Respiration ◽  
Phosphoglycerate Kinase ◽  
Anaerobic Growth ◽  
Wild Type ◽  
Kinase Gene ◽  
Glycolytic Intermediate

ABSTRACT The Bacillus subtilis ResD-ResE two-component signal transduction system is essential for aerobic and anaerobic respiration. A spontaneous suppressor mutant that expresses ResD-controlled genes and grows anaerobically in the absence of the ResE histidine kinase was isolated. In addition, aerobic expression of ResD-controlled genes in the suppressed strain was constitutive and occurred at a much higher level than that observed in the wild-type strain. The suppressing mutation, which mapped to pgk, the gene encoding 3-phosphoglycerate kinase, failed to suppress a resDmutation, suggesting that the suppressing mutation creates a pathway for phosphorylation of the response regulator, ResD, which is independent of the cognate sensor kinase, ResE. The pgk-1mutant exhibited very low but measurable 3-phosphoglycerate kinase activity compared to the wild-type strain. The results suggest that accumulation of a glycolytic intermediate, probably 1,3-diphosphoglycerate, is responsible for the observed effect of thepgk-1 mutation on anaerobiosis of resE mutant cells.

scholarly journals Regulation and Characterization of a Newly Deduced Cell Wall Hydrolase Gene (cwlJ) Which Affects Germination of Bacillus subtilis Spores

1998 ◽  
Vol 180 (6) ◽  
pp. 1375-1380 ◽  
Author(s):  
Shu Ishikawa ◽  
Kunio Yamane ◽  
Junichi Sekiguchi
Keyword(s):  
Bacillus Subtilis ◽  
Type Strain ◽  
Wild Type Strain ◽  
Catalytic Domain ◽  
Slow Release ◽  
Dipicolinic Acid ◽  
Northern Hybridization ◽  
Predicted Amino Acid Sequence ◽  
Wild Type

ABSTRACT The predicted amino acid sequence of Bacillus subtilis ycbQ (renamed cwlJ) exhibits high similarity to those of the deduced C-terminal catalytic domain of SleBs, the specific cortex-hydrolyzing enzyme of B. cereus and the deduced one of B. subtilis. We constructed acwlJ::lacZ fusion in the B. subtilischromosome. The β-galactosidase activity and results of Northern hybridization and primer extension analyses of the cwlJgene indicated that it is transcribed by EςE RNA polymerase. cwlJ-deficient spores responded to bothl-alanine and AGFK, the A 580 values of spore suspensions decreased more slowly than in the case of the wild-type strain, and the mutant spores released less dipicolinic acid than did those of the wild-type strain during germination. However, the mutant spores released only slightly less hexosamine than did the wild-type spores. In contrast, B. subtilis sleB spores did not release hexosamine at a significant level. While cwlJand sleB spores were able to germinate, CJSB (cwlJ sleB) spores could not germinate but exhibited initial germination reactions, e.g., partial decrease inA 580 and slow release of dipicolinic acid. CJSB spores became slightly gray after 6 h in the germinant, but their refractility was much greater than that of sleB mutant spores. The roles of the sleB and cwlJmutations in germination and spore maturation are also discussed.

scholarly journals The dlt Operon of Bacillus cereus Is Required for Resistance to Cationic Antimicrobial Peptides and for Virulence in Insects

2009 ◽  
Vol 191 (22) ◽  
pp. 7063-7073 ◽  
Author(s):  
Z. Abi Khattar ◽  
A. Rejasse ◽  
D. Destoumieux-Garzón ◽  
J. M. Escoubas ◽  
V. Sanchis ◽  
...  
Keyword(s):  
Antimicrobial Peptides ◽  
Bacillus Cereus ◽  
Cell Walls ◽  
Type Strain ◽  
Pathogenic Bacteria ◽  
Wild Type Strain ◽  
Wild Type ◽  
Gram Positive ◽  
Cationic Antimicrobial Peptides ◽  
Humoral Immune

ABSTRACT The dlt operon encodes proteins that alanylate teichoic acids, the major components of cell walls of gram-positive bacteria. This generates a net positive charge on bacterial cell walls, repulsing positively charged molecules and conferring resistance to animal and human cationic antimicrobial peptides (AMPs) in gram-positive pathogenic bacteria. AMPs damage the bacterial membrane and are the most effective components of the humoral immune response against bacteria. We investigated the role of the dlt operon in insect virulence by inactivating this operon in Bacillus cereus, which is both an opportunistic human pathogen and an insect pathogen. The ΔdltBc mutant displayed several morphological alterations but grew at a rate similar to that for the wild-type strain. This mutant was less resistant to protamine and several bacterial cationic AMPs, such as nisin, polymyxin B, and colistin, in vitro. It was also less resistant to molecules from the insect humoral immune system, lysozyme, and cationic AMP cecropin B from Spodoptera frugiperda. ΔdltBc was as pathogenic as the wild-type strain in oral infections of Galleria mellonella but much less virulent when injected into the hemocoels of G. mellonella and Spodoptera littoralis. We detected the dlt operon in three gram-negative genera: Erwinia (Erwinia carotovora), Bordetella (Bordetella pertussis, Bordetella parapertussis, and Bordetella bronchiseptica), and Photorhabdus (the entomopathogenic bacterium Photorhabdus luminescens TT01, the dlt operon of which did not restore cationic AMP resistance in ΔdltBc ). We suggest that the dlt operon protects B. cereus against insect humoral immune mediators, including hemolymph cationic AMPs, and may be critical for the establishment of lethal septicemia in insects and in nosocomial infections in humans.

scholarly journals Bacillus subtilis Phosphorylated PhoP: Direct Activation of the EσA- and Repression of the EσE-Responsive phoB-PS+V Promoters during Pho Response

2005 ◽  
Vol 187 (15) ◽  
pp. 5166-5178 ◽  
Author(s):  
Wael R. Abdel-Fattah ◽  
Yinghua Chen ◽  
Amr Eldakak ◽  
F. Marion Hulett
Keyword(s):  
Alkaline Phosphatase ◽  
Bacillus Subtilis ◽  
Type Strain ◽  
Wild Type Strain ◽  
Pho Regulon ◽  
Dependent Manner ◽  
Wild Type ◽  
Phosphate Deprivation

ABSTRACT The phoB gene of Bacillus subtilis encodes an alkaline phosphatase (PhoB, formerly alkaline phosphatase III) that is expressed from separate promoters during phosphate deprivation in a PhoP-PhoR-dependent manner and at stage two of sporulation under phosphate-sufficient conditions independent of PhoP-PhoR. Isogenic strains containing either the complete phoB promoter or individual phoB promoter fusions were used to assess expression from each promoter under both induction conditions. The phoB promoter responsible for expression during sporulation, phoB-PS, was expressed in a wild-type strain during phosphate deprivation, but induction occurred >3 h later than induction of Pho regulon genes and the levels were approximately 50-fold lower than that observed for the PhoPR-dependent promoter, phoB-PV. EσE was necessary and sufficient for PS expression in vitro. PS expression in a phoPR mutant strain was delayed 2 to 3 h compared to the expression in a wild-type strain, suggesting that expression or activation of σE is delayed in a phoPR mutant under phosphate-deficient conditions, an observation consistent with a role for PhoPR in spore development under these conditions. Phosphorylated PhoP (PhoP∼P) repressed PS in vitro via direct binding to the promoter, the first example of an EσE-responsive promoter that is repressed by PhoP∼P. Whereas either PhoP or PhoP∼P in the presence of EσA was sufficient to stimulate transcription from the phoB-PV promoter in vitro, roughly 10- and 17-fold-higher concentrations of PhoP than of PhoP∼P were required for PV promoter activation and maximal promoter activity, respectively. The promoter for a second gene in the Pho regulon, ykoL, was also activated by elevated concentrations of unphosphorylated PhoP in vitro. However, because no Pho regulon gene expression was observed in vivo during Pi -replete growth and PhoP concentrations increased only threefold in vivo during phoPR autoinduction, a role for unphosphorylated PhoP in Pho regulon activation in vivo is not likely.

scholarly journals Transcription of the Bacillus subtilis gerK Operon, Which Encodes a Spore Germinant Receptor, and Comparison with That of Operons Encoding Other Germinant Receptors

2006 ◽  
Vol 188 (11) ◽  
pp. 4131-4136 ◽  
Author(s):  
Takao Igarashi ◽  
Peter Setlow
Keyword(s):  
Bacillus Subtilis ◽  
Sigma Factor ◽  
Type Strain ◽  
Wild Type Strain ◽  
Dipicolinic Acid ◽  
Mrna Levels ◽  
Wild Type ◽  
Putative Promoter ◽  
Rapid Fall ◽  
Sequence Similarities

ABSTRACT The gerA, gerB, and gerK operons, which encode germinant receptors in spores of Bacillus subtilis, were transcribed only in sporulation, and their mRNA levels peaked initially ∼3 h before the initiation of accumulation of the spore's dipicolinic acid. After a rapid fall, levels of these mRNAs peaked again ∼5 h later. In one wild-type strain (PS832), gerA mRNA was the most abundant, with levels of gerB and gerK mRNAs ∼50% of that of gerA mRNA, whereas gerB mRNA was the most abundant in another wild-type strain (PY79). The synthesis of gerK mRNA in sporulation was abolished by loss of the forespore-specific RNA polymerase sigma factor, σG, and induction of σG synthesis in vegetative cells led to synthesis of gerK mRNA. SpoVT, a regulator of σG-dependent gene expression, repressed gerK expression. The gerK promoter showed sequence similarities to σG-dependent promoters, and deletion of elements of this putative promoter abolished gerK expression in sporulation.

scholarly journals Genetic Analysis of an Incomplete mutSGene from Pseudomonas putida

2000 ◽  
Vol 182 (18) ◽  
pp. 5278-5279 ◽  
Author(s):  
Yasurou Kurusu ◽  
Tomoaki Narita ◽  
Makoto Suzuki ◽  
Taeko Watanabe
Keyword(s):  
Escherichia Coli ◽  
Bacillus Subtilis ◽  
Genetic Analysis ◽  
Pseudomonas Putida ◽  
Type Strain ◽  
Mutation Frequency ◽  
Wild Type Strain ◽  
Wild Type

ABSTRACT We genetically characterized the Pseudomonas putida mutS gene and found that it encodes a smaller MutS protein than do the genes of other bacteria. This gene is able to function in themutS mutants of Escherichia coli andBacillus subtilis. A P. putida mutS mutant has a mutation frequency 1,000-fold greater than that of the wild-type strain.

scholarly journals Controlling the hypermutation: Exploitation of the MutS protein in Pseudomonas aeruginosa

2020 ◽  
Vol 2 (7A) ◽  
Author(s):  
Yue Yuan On ◽  
Martin Welch
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Type Strain ◽  
Wild Type Strain ◽  
Resistant Mutant ◽  
Inducible Promoter ◽  
Wild Type ◽  
Cdna Analysis ◽  
In The Wild ◽  
Mismatch Recognition ◽  
Muts Gene

Pseudomonas aeruginosa infections commonly develop in individuals with cystic fibrosis (CF), and its adaptation in such an unfavourable condition is always found to be related to hypermutation. In fact, most of the hypermutation is due to the defects in mutS gene which involves in the mismatch repair mechanism, causing the acceleration of mutation rate and adaptive evolution. In order to rheostatically express the MutS protein and achieve “hypomutation” (in which the rate of mutation is lower than that of wild type strain), an exogenous mutS gene with rhamnose-inducible promoter was cloned into MPAO1 mutS::Tn mutant strain. Present findings demonstrate that this system is tightly-controlled and stable, with less rifampicin-resistant mutant frequency and more fluorescence intensity from a GFP-tagged MutS expressing cells were observed when the concentration of the inducer increases. Interestingly, the results from Western blot analysis show that less MutS protein is required to suppress hypermutation in the wild type strain, as compared to our construct that behaves similar to the wild type but obviously needs more MutS expression to achieve such state. This indicates that the exogenous MutS might be lacking of other important protein to work efficiently in mismatch recognition. Therefore, based on our cDNA analysis, we found that fdxA gene next to the mutS gene is in the same operon, which could suggest that they might be functionally related in the DNA repair machinery.

Sign in / Sign up

Export Citation Format

Share Document