scholarly journals Biochemical Genetics of the Cryptic Gene System for Cellobiose Utilization in Escherichia coli K12

Genetics ◽  
1987 ◽  
Vol 115 (3) ◽  
pp. 419-429
Author(s):  
Maja Kricker ◽  
Barry G Hall
Keyword(s):  
Escherichia Coli ◽  
Microbial Evolution ◽  
Transport Protein ◽  
Wild Type ◽  
Phosphotransferase System ◽  
E Coli ◽  
Escherichia Coli K12 ◽  
Biochemical Assays ◽  
In The Wild

ABSTRACT The cellobiose catabolic system of Escherichia coli K12 is being used to study the role of cryptic genes in microbial evolution. Wild-type E. coli K12 do not utilize the β-glucoside sugars, arbutin, salicin and cellobiose. A Cel+ (cellobiose utilizing) mutant which grows on cellobiose, arbutin, and salicin was isolated previously from wild-type E. coli K12. Biochemical assays indicate that a cel structural gene (celT) specifies a single transport protein that is a β-glucoside specific enzyme of the phosphoenolpyruvate-dependent phosphotransferase system. The transport protein phosphorylates β-glucosides at the expense of phosphoenolpyruvate. A single phosphoglucosidase, specified by celH, hydrolyzes phosphorylated cellobiose, arbutin, and salicin. The genes of the cel system are expressed constitutively in the Cel+ mutant, whereas they are not expressed at a detectable level in the wild-type strain. The transport and hydrolase genes are simultaneously silenced or simultaneously expressed and thus constitute an operon. Cel+ strains which fail to utilize one or more β-glucosides express the transport system at a lower level than do Cel+ strains which grow on all three β-glucosides. Other strains inducibly express a gene which specifies transport of arbutin but not the other β-glucosides. The arbutin transport gene, arbT, maps outside of the cel locus.

Isolation and characterization of catabolite repression-resistant mutants of Escherichia coli

1977 ◽  
Vol 23 (10) ◽  
pp. 1384-1393 ◽  
Author(s):  
Glen D. Armstrong ◽  
Hiroshi Yamazaki
Keyword(s):  
Escherichia Coli ◽  
Catabolite Repression ◽  
Coli Strain ◽  
Wild Type ◽  
Phosphotransferase System ◽  
E Coli ◽  
Isolation And Characterization ◽  
In The Wild ◽  
Resistant Mutants

A method has been developed for the isolation of Escherichia coli mutants which are resistant to catabolite repression. The method is based on the fact that a mixture of glucose and gluconate inhibits the development of chemotactic motility in the wild type, but not in the mutants. A motile E. coli strain was mutagenized and grown in glucose and gluconate. Mutants which were able to swim into a tube containing a chemotactic attractant (aspartic acid) were isolated. Most of these mutants were able to produce β-galactosidase in the presence of glucose and gluconate and were normal in their ability to degrade adenosine 3′,5′-cyclic monophosphate. Some of these mutants were defective in the glucose phosphotransferase system.

O6-methylguanine-DNA methyltransferase in wild-type and ada mutants of Escherichia coli

1982 ◽  
Vol 152 (1) ◽  
pp. 534-537
Author(s):  
S Mitra ◽  
B C Pal ◽  
R S Foote
Keyword(s):  
Escherichia Coli ◽  
Dna Methyltransferase ◽  
Wild Type ◽  
E Coli ◽  
Methylguanine Dna Methyltransferase ◽  
Synthetic Dna ◽  
In The Wild ◽  
Low Levels ◽  
Enzyme Molecules ◽  
Dna Substrate

O(6)-Methylguanine-DNA methyltransferase is induced in Escherichia coli during growth in low levels of N-methyl-N'-nitro-N-nitrosoguanidine. We have developed a sensitive assay for quantitating low levels of this activity with a synthetic DNA substrate containing 3H-labeled O(6)-methylguanine as the only modified base. Although both wild-type and adaptation-deficient (ada) mutants of E. coli contained low but comparable numbers (from 13 to 60) of the enzyme molecules per cell, adaptation treatment caused a significant increase of the enzyme in the wild type but not in the ada mutants, suggesting that the ada mutation is in a regulatory locus and not in the structural gene for the methyltransferase.

scholarly journals Transposon Insertion in the purL Gene Induces Biofilm Depletion in Escherichia coli ATCC 25922

Pathogens ◽  
2020 ◽  
Vol 9 (9) ◽  
pp. 774
Author(s):  
Virginio Cepas ◽  
Victoria Ballén ◽  
Yaiza Gabasa ◽  
Miriam Ramírez ◽  
Yuly López ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Biofilm Formation ◽  
De Novo ◽  
Wild Type ◽  
Planktonic Cells ◽  
E Coli ◽  
Qrt Pcr ◽  
Transposon Insertion ◽  
De Novo Purine Biosynthesis

Current Escherichia coli antibiofilm treatments comprise a combination of antibiotics commonly used against planktonic cells, leading to treatment failure. A better understanding of the genes involved in biofilm formation could facilitate the development of efficient and specific new antibiofilm treatments. A total of 2578 E. coli mutants were generated by transposon insertion, of which 536 were analysed in this study. After sequencing, Tn263 mutant, classified as low biofilm-former (LF) compared to the wild-type (wt) strain (ATCC 25922), showed an interruption in the purL gene, involved in the de novo purine biosynthesis pathway. To elucidate the role of purL in biofilm formation, a knockout was generated showing reduced production of curli fibres, leading to an impaired biofilm formation. These conditions were restored by complementation of the strain or addition of exogenous inosine. Proteomic and transcriptional analyses were performed to characterise the differences caused by purL alterations. Thirteen proteins were altered compared to wt. The corresponding genes were analysed by qRT-PCR not only in the Tn263 and wt, but also in clinical strains with different biofilm activity. Overall, this study suggests that purL is essential for biofilm formation in E. coli and can be considered as a potential antibiofilm target.

scholarly journals The RcsCB His-Asp Phosphorelay System Is Essential To Overcome Chlorpromazine-Induced Stress in Escherichia coli

2002 ◽  
Vol 184 (10) ◽  
pp. 2850-2853 ◽  
Author(s):  
Annie Conter ◽  
Rachel Sturny ◽  
Claude Gutierrez ◽  
Kaymeuang Cam
Keyword(s):  
Escherichia Coli ◽  
Type Strain ◽  
Wild Type Strain ◽  
Cell Envelope ◽  
Wild Type ◽  
E Coli ◽  
Induced Stress ◽  
Mutant Strains ◽  
Molecular Nature

ABSTRACT The RcsCB His-Asp phosphorelay system regulates the expression of several genes of Escherichia coli, but the molecular nature of the inducing signal is still unknown. We show here that treatment of an exponentially growing culture of E. coli with the cationic amphipathic compound chlorpromazine (CPZ) stimulates expression of a set of genes positively regulated by the RcsCB system. This induction is abolished in rcsB or rcsC mutant strains. In addition, treatment with CPZ inhibits growth. The wild-type strain is able to recover from this inhibition and resume growth after a period of adaptation. In contrast, strains deficient in the RcsCB His-Asp phosphorelay system are hypersensitive to CPZ. These results suggest that cells must express specific RcsCB-regulated genes in order to cope with the CPZ-induced stress. This is the first report of the essential role of the RcsCB system in a stress situation. These results also strengthen the notion that alterations of the cell envelope induce a signal recognized by the RcsC sensor.

scholarly journals Role of Motility and the flhDC Operon in Escherichia coli MG1655 Colonization of the Mouse Intestine

2007 ◽  
Vol 75 (7) ◽  
pp. 3315-3324 ◽  
Author(s):  
Eric J. Gauger ◽  
Mary P. Leatham ◽  
Regino Mercado-Lubo ◽  
David C. Laux ◽  
Tyrrell Conway ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Regulatory Region ◽  
Flagellar Motor ◽  
Wild Type ◽  
E Coli ◽  
Mouse Intestine ◽  
Flagellum Biosynthesis ◽  
Colonizing Ability

ABSTRACT Previously, we reported that the mouse intestine selected mutants of Escherichia coli MG1655 that have improved colonizing ability (M. P. Leatham et al., Infect. Immun. 73:8039-8049, 2005). These mutants grew 10 to 20% faster than their parent in mouse cecal mucus in vitro and 15 to 30% faster on several sugars found in the mouse intestine. The mutants were nonmotile and had deletions of various lengths beginning immediately downstream of an IS1 element located within the regulatory region of the flhDC operon, which encodes the master regulator of flagellum biosynthesis, FlhD4C2. Here we show that during intestinal colonization by wild-type E. coli strain MG1655, 45 to 50% of the cells became nonmotile by day 3 after feeding of the strain to mice and between 80 and 90% of the cells were nonmotile by day 15 after feeding. Ten nonmotile mutants isolated from mice were sequenced, and all were found to have flhDC deletions of various lengths. Despite this strong selection, 10 to 20% of the E. coli MG1655 cells remained motile over a 15-day period, suggesting that there is an as-yet-undefined intestinal niche in which motility is an advantage. The deletions appear to be selected in the intestine for two reasons. First, genes unrelated to motility that are normally either directly or indirectly repressed by FlhD4C2 but can contribute to maximum colonizing ability are released from repression. Second, energy normally used to synthesize flagella and turn the flagellar motor is redirected to growth.

scholarly journals Role of CheB and CheR in the Complex Chemotactic and Aerotactic Pathway of Azospirillum brasilense

2006 ◽  
Vol 188 (13) ◽  
pp. 4759-4768 ◽  
Author(s):  
Bonnie B. Stephens ◽  
Star N. Loar ◽  
Gladys Alexandre
Keyword(s):  
Escherichia Coli ◽  
Steady State ◽  
Azospirillum Brasilense ◽  
Model Organism ◽  
Spatial Gradient ◽  
Wild Type ◽  
Temporal Gradient ◽  
E Coli ◽  
Significant Difference

ABSTRACT It has previously been reported that the alpha-proteobacterium Azospirillum brasilense undergoes methylation-independent chemotaxis; however, a recent study revealed cheB and cheR genes in this organism. We have constructed cheB, cheR, and cheBR mutants of A. brasilense and determined that the CheB and CheR proteins under study significantly influence chemotaxis and aerotaxis but are not essential for these behaviors to occur. First, we found that although cells lacking CheB, CheR, or both were no longer capable of responding to the addition of most chemoattractants in a temporal gradient assay, they did show a chemotactic response (albeit reduced) in a spatial gradient assay. Second, in comparison to the wild type, cheB and cheR mutants under steady-state conditions exhibited an altered swimming bias, whereas the cheBR mutant and the che operon mutant did not. Third, cheB and cheR mutants were null for aerotaxis, whereas the cheBR mutant showed reduced aerotaxis. In contrast to the swimming bias for the model organism Escherichia coli, the swimming bias in A. brasilense cells was dependent on the carbon source present and cells released methanol upon addition of some attractants and upon removal of other attractants. In comparison to the wild type, the cheB, cheR, and cheBR mutants showed various altered patterns of methanol release upon exposure to attractants. This study reveals a significant difference between the chemotaxis adaptation system of A. brasilense and that of the model organism E. coli and suggests that multiple chemotaxis systems are present and contribute to chemotaxis and aerotaxis in A. brasilense.

scholarly journals Role of Quorum Sensing and Antimicrobial Component Production by Serratia plymuthica in Formation of Biofilms, Including Mixed Biofilms with Escherichia coli

2006 ◽  
Vol 72 (11) ◽  
pp. 7294-7300 ◽  
Author(s):  
Pieter Moons ◽  
Rob Van Houdt ◽  
Abram Aertsen ◽  
Kristof Vanoirbeek ◽  
Yves Engelborghs ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Quorum Sensing ◽  
Biofilm Formation ◽  
Homoserine Lactone ◽  
Broth Culture ◽  
Confocal Laser ◽  
Wild Type ◽  
Serratia Plymuthica ◽  
E Coli

ABSTRACT We have previously characterized the N-acyl-l-homoserine lactone-based quorum-sensing system of the biofilm isolate Serratia plymuthica RVH1. Here we investigated the role of quorum sensing and of quorum-sensing-dependent production of an antimicrobial compound (AC) on biofilm formation by RVH1 and on the cocultivation of RVH1 and Escherichia coli in planktonic cultures or in biofilms. Biofilm formation of S. plymuthica was not affected by the knockout of splI or splR, the S. plymuthica homologs of the luxI or luxR quorum-sensing gene, respectively, or by the knockout of AC production. E. coli grew well in mixed broth culture with RVH1 until the latter reached 8.5 to 9.5 log CFU/ml, after which the E. coli colony counts steeply declined. In comparison, only a very small decline occurred in cocultures with the S. plymuthica AC-deficient and splI mutants. Complementation with exogenous N-hexanoyl-l-homoserine lactone rescued the wild-type phenotype of the splI mutant. The splR knockout mutant also induced a steep decline of E. coli, consistent with its proposed function as a repressor of quorum-sensing-regulated genes. The numbers of E. coli in 3-day-old mixed biofilms followed a similar pattern, being higher with S. plymuthica deficient in SplI or AC production than with wild-type S. plymuthica, the splR mutant, or the splI mutant in the presence of N-hexanoyl-l-homoserine lactone. Confocal laser scanning microscopic analysis of mixed biofilms established with strains producing different fluorescent proteins showed that E. coli microcolonies were less developed in the presence of RVH1 than in the presence of the AC-deficient mutant.

scholarly journals Identification of a Phosphotransferase System of Escherichia coli Required for Growth on N-Acetylmuramic Acid

2004 ◽  
Vol 186 (8) ◽  
pp. 2385-2392 ◽  
Author(s):  
Ulrike Dahl ◽  
Tina Jaeger ◽  
Bao Trâm Nguyen ◽  
Julia M. Sattler ◽  
Christoph Mayer
Keyword(s):  
Escherichia Coli ◽  
Energy Analysis ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Sole Source ◽  
Wild Type ◽  
Phosphotransferase System ◽  
E Coli ◽  
Glucose Transport System ◽  
Single Polypeptide

ABSTRACT We report here that wild-type Escherichia coli grows on N-acetylmuramic acid (MurNAc) as the sole source of carbon and energy. Analysis of mutants defective in N-acetylglucosamine (GlcNAc) catabolism revealed that the catabolic pathway for MurNAc merges into the GlcNAc pathway on the level of GlcNAc 6-phosphate. Furthermore, analysis of mutants defective in components of the phosphotransferase system (PTS) revealed that a PTS is essential for growth on MurNAc. However, neither the glucose-, mannose/glucosamine-, nor GlcNAc-specific PTS (PtsG, ManXYZ, and NagE, respectively) was found to be necessary. Instead, we identified a gene at 55 min on the E. coli chromosome that is responsible for MurNAc uptake and growth. It encodes a single polypeptide consisting of the EIIB and C domains of a so-far-uncharacterized PTS that was named murP. MurP lacks an EIIA domain and was found to require the activity of the crr-encoded enzyme IIA-glucose (EIIAGlc), a component of the major glucose transport system for growth on MurNAc. murP deletion mutants were unable to grow on MurNAc as the sole source of carbon; however, growth was rescued by providing murP in trans expressed from an isopropylthiogalactopyranoside-inducible plasmid. A functional His6 fusion of MurP was constructed, isolated from membranes, and identified as a polypeptide with an apparent molecular mass of 37 kDa by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and Western blot analysis. Close homologs of MurP were identified in the genome of several bacteria, and we believe that these organisms might also be able to utilize MurNAc.

scholarly journals Substrate Specificity and Signal Transduction Pathways in the Glucose-Specific Enzyme II (EIIGlc) Component of the Escherichia coli Phosphotransferase System

2000 ◽  
Vol 182 (16) ◽  
pp. 4437-4442 ◽  
Author(s):  
Lucinda Notley-McRobb ◽  
Thomas Ferenci
Keyword(s):  
Escherichia Coli ◽  
Signal Transduction ◽  
Substrate Specificity ◽  
Dependent Manner ◽  
Wild Type ◽  
Specific Enzyme ◽  
Phosphotransferase System ◽  
Transmembrane Segments ◽  
In The Wild ◽  
Second Category

ABSTRACT Escherichia coli adapted to glucose-limited chemostats contained mutations in ptsG resulting in V12G, V12F, and G13C substitutions in glucose-specific enzyme II (EIIGlc) and resulting in increased transport of glucose and methyl-α-glucoside. The mutations also resulted in faster growth on mannose and glucosamine in a PtsG-dependent manner. By use of enhanced growth on glucosamine for selection, four further sites were identified where substitutions caused broadened substrate specificity (G176D, A288V, G320S, and P384R). The altered amino acids include residues previously identified as changing the uptake of ribose, fructose, and mannitol. The mutations belonged to two classes. First, at two sites, changes affected transmembrane residues (A288V and G320S), probably altering sugar selectivity directly. More remarkably, the five other specificity mutations affected residues unlikely to be in transmembrane segments and were additionally associated with increasedptsG transcription in the absence of glucose. Increased expression of wild-type EIIGlc was not by itself sufficient for growth with other sugars. A model is proposed in which the protein conformation determining sugar accessibility is linked to transcriptional signal transduction in EIIGlc. The conformation of EIIGlc elicited by either glucose transport in the wild-type protein or permanently altered conformation in the second category of mutants results in altered signal transduction and interaction with a regulator, probably Mlc, controlling the transcription of pts genes.

Biofilm Growth of Escherichia coli Is Subject to cAMP-Dependent and cAMP-Independent Inhibition

2015 ◽  
Vol 25 (2-3) ◽  
pp. 209-225 ◽  
Author(s):  
Sarah L. Sutrina ◽  
Kia Daniel ◽  
Michael Lewis ◽  
Naomi T. Charles ◽  
Cherysa K.E. Anselm ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Catabolite Repression ◽  
Ph Effects ◽  
Wild Type ◽  
Inhibitory Effects ◽  
Phosphotransferase System ◽  
Biofilm Growth ◽  
E Coli ◽  
Low Concentrations ◽  
High Concentrations

We established that <i>Escherichia coli </i>strain 15 (ATCC 9723) produces both curli and cellulose, and forms robust biofilms. Since this strain is wild type with respect to the phosphoenolpyruvate:sugar phosphotransferase system (PTS), it is an ideal strain in which to investigate the effects of the PTS on the biofilm growth of <i>E. coli</i>. We began by looking into the effects of PTS and non-PTS sugars on the biofilm growth of this strain. All the sugars tested tended to activate biofilm growth at low concentrations but to inhibit biofilm growth at high concentrations. Acidification of the medium was an inhibitory factor in the absence of buffer, but buffering to prevent a pH drop did not prevent the inhibitory effects of the sugars. The concentration at which inhibition set in varied from sugar to sugar. For most sugars, cyclic (c)AMP counteracted the inhibition at the lowest inhibitory concentrations but became ineffective at higher concentrations. Our results suggest that cAMP-dependent catabolite repression, which is mediated by the PTS in <i>E. coli</i>, plays a role in the regulation of biofilm growth in response to sugars. cAMP-independent processes, possibly including Cra, also appear to be involved, in addition to pH effects.

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