scholarly journals Role of Ribosome Release in Regulation oftna Operon Expression in Escherichia coli

1999 ◽  
Vol 181 (5) ◽  
pp. 1530-1536 ◽  
Author(s):  
Kouacou Vincent Konan ◽  
Charles Yanofsky
Keyword(s):  
Escherichia Coli ◽  
Mutant Allele ◽  
Stop Codon ◽  
Leader Peptide ◽  
Coding Region ◽  
Reporter Construct ◽  
E Coli ◽  
Operon Expression ◽  
Basal Level Expression

ABSTRACT Expression of the degradative tryptophanase (tna) operon of Escherichia coli is regulated by catabolite repression and tryptophan-induced transcription antitermination. In cultures growing in the absence of added tryptophan, transcription of the structural genes of the tna operon is limited by Rho-dependent transcription termination in the leader region of the operon. Tryptophan induction prevents this Rho-dependent termination, and requires in-frame translation of a 24-residue leader peptide coding region, tnaC, that contains a single, crucial, Trp codon. Studies with a lacZ reporter construct lacking the spacer region between tnaC and the first major structural gene,tnaA, suggested that tryptophan induction might involvecis action by the TnaC leader peptide on the ribosome translating the tnaC coding region. The leader peptide was hypothesized to inhibit ribosome release at thetnaC stop codon, thereby blocking Rho’s access to the transcript. Regulatory studies with deletion constructs of thetna operon of Proteus vulgaris supported this interpretation. In the present study the putative role of thetnaC stop codon in tna operon regulation inE. coli was examined further by replacing the naturaltnaC stop codon, UGA, with UAG or UAA in atnaC-stop codon-tnaA′-′lacZ reporter construct. Basal level expression was reduced to 20 and 50% when the UGA stop codon was replaced by UAG or UAA, respectively, consistent with the finding that in E. coli translation terminates more efficiently at UAG and UAA than at UGA. Tryptophan induction was observed in strains with any of the stop codons. However, when UAG or UAA replaced UGA, the induced level of expression was also reduced to 15 and 50% of that obtained with UGA as the tnaC stop codon, respectively. Introduction of a mutant allele encoding a temperature-sensitive release factor 1, prfA1, increased basal level expression 60-fold when the tnaC stop codon was UAG and 3-fold when this stop codon was UAA; basal level expression was reduced by 50% in the construct with the natural stop codon, UGA. In strains with any of the three stop codons and the prfA1mutation, the induced levels of tna operon expression were virtually identical. The effects of tnaC stop codon identity on expression were also examined in the absence of Rho action, using tnaC-stop codon-′lacZ constructs that lack the tnaC-tnaA spacer region. Expression was low in the absence of tnaC stop codon suppression. In most cases, tryptophan addition resulted in about 50% inhibition of expression when UGA was replaced by UAG or UAA and the appropriate suppressor was present. Introduction of the prfA1 mutant allele increased expression of the suppressed construct with the UAG stop codon; tryptophan addition also resulted in ca. 50% inhibition. These findings provide additional evidence implicating the behavior of the ribosome translating tnaC in the regulation of tna operon expression.

scholarly journals Identification of Escherichia coli ubiB, a Gene Required for the First Monooxygenase Step in Ubiquinone Biosynthesis

2000 ◽  
Vol 182 (18) ◽  
pp. 5139-5146 ◽  
Author(s):  
Wayne W. Poon ◽  
Diana E. Davis ◽  
Huan T. Ha ◽  
Tanya Jonassen ◽  
Philip N. Rather ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Saccharomyces Cerevisiae ◽  
Protein Kinase ◽  
Mutant Allele ◽  
Coenzyme Q ◽  
Low Frequency ◽  
Coding Region ◽  
E Coli ◽  
Mutant Strains ◽  
Providencia Stuartii

ABSTRACT It was recently discovered that the aarF gene inProvidencia stuartii is required for coenzyme Q (CoQ) biosynthesis. Here we report that yigR, theEscherichia coli homologue of aarF, isubiB, a gene required for the first monooxygenase step in CoQ biosynthesis. Both the P. stuartii aarF and E. coli ubiB (yigR) disruption mutant strains lack CoQ and accumulate octaprenylphenol. Octaprenylphenol is the CoQ biosynthetic intermediate found to accumulate in the E. coli strain AN59, which contains the ubiB409 mutant allele. Analysis of the mutation in the E. coli strain AN59 reveals no mutations within the ubiB gene, but instead shows the presence of an IS1 element at position +516 of the ubiE gene. The ubiE gene encodes aC-methyltransferase required for the synthesis of both CoQ and menaquinone, and it is the 5′ gene in an operon containingubiE, yigP, and ubiB. The data indicate that octaprenylphenol accumulates in AN59 as a result of a polar effect of the ubiE::IS1mutation on the downstream ubiB gene. AN59 is complemented by a DNA segment containing the contiguous ubiE,yigP, and ubiB genes. Although transformation of AN59 with a DNA segment containing the ubiB coding region fails to restore CoQ biosynthesis, transformation with theubiE coding region results in a low-frequency but significant rescue attributed to homologous recombination. In addition, the fre gene, previously considered to correspond toubiB, was found not to be involved in CoQ biosynthesis. TheubiB gene is a member of a predicted protein kinase family of which the Saccharomyces cerevisiae ABC1 gene is the prototypic member. The possible protein kinase function of UbiB and Abc1 and the role these polypeptides may play in CoQ biosynthesis are discussed.

scholarly journals Rho-Dependent Transcription Termination in the tnaOperon of Escherichia coli: Roles of the boxASequence and the rut Site

2000 ◽  
Vol 182 (14) ◽  
pp. 3981-3988 ◽  
Author(s):  
Kouacou Vincent Konan ◽  
Charles Yanofsky
Keyword(s):  
Escherichia Coli ◽  
Stop Codon ◽  
Transcription Termination ◽  
Point Mutations ◽  
Constitutive Expression ◽  
Peptide Sequence ◽  
Start Codon ◽  
Leader Peptide ◽  
Coding Region ◽  
High Level

ABSTRACT Expression of the tryptophanase (tna) operon ofEscherichia coli is regulated by catabolite repression and by tryptophan-induced transcription antitermination. Tryptophan induction prevents Rho-dependent transcription termination in the leader region of the operon. Induction requires translation of a 24-residue leader peptide-coding region, tnaC, containing a single, crucial Trp codon. Studies with a lacZ reporter construct lacking the tnaC-tnaA spacer region suggest that, in the presence of excess tryptophan, the TnaC leader peptide acts incis on the ribosome translating tnaC to inhibit its release. The stalled ribosome is thought to block Rho's access to the transcript. In this paper we examine the roles of theboxA sequence and the rut site in Rho-dependent termination. Deleting six nucleotides (CGC CCT) of boxA or introducing specific point mutations in boxA results in high-level constitutive expression. Some constitutive changes introduced in boxA do not change the TnaC peptide sequence. We confirm that deletion of the rut site results in constitutive expression. We also demonstrate that, in each constitutive construct, replacement of the tnaC start codon by a UAG stop codon reduces expression significantly, suggesting that constitutive expression requires translation of the tnaCcoding sequence. Addition of bicyclomycin, an inhibitor of Rho, to these UAG constructs increases expression, demonstrating that reduced expression is due to Rho action. Combining a boxA point mutation with rut site deletion results in constitutive expression comparable to that of a maximally induced operon. These results support the hypothesis that in the presence of tryptophan the ribosome translating tnaC blocks Rho's access to theboxA and rut sites, thereby preventing transcription termination.

scholarly journals Roles of the C-Terminal Amino Acids of Non-Hexameric Helicases: Insights from Escherichia coli UvrD

2021 ◽  
Vol 22 (3) ◽  
pp. 1018
Author(s):  
Hiroaki Yokota
Keyword(s):  
Escherichia Coli ◽  
Amino Acids ◽  
Amino Acid ◽  
Single Molecule ◽  
Underlying Mechanism ◽  
Amino Acid Sequences ◽  
E Coli ◽  
X Ray Crystallography ◽  
Terminal Amino

Helicases are nucleic acid-unwinding enzymes that are involved in the maintenance of genome integrity. Several parts of the amino acid sequences of helicases are very similar, and these quite well-conserved amino acid sequences are termed “helicase motifs”. Previous studies by X-ray crystallography and single-molecule measurements have suggested a common underlying mechanism for their function. These studies indicate the role of the helicase motifs in unwinding nucleic acids. In contrast, the sequence and length of the C-terminal amino acids of helicases are highly variable. In this paper, I review past and recent studies that proposed helicase mechanisms and studies that investigated the roles of the C-terminal amino acids on helicase and dimerization activities, primarily on the non-hexermeric Escherichia coli (E. coli) UvrD helicase. Then, I center on my recent study of single-molecule direct visualization of a UvrD mutant lacking the C-terminal 40 amino acids (UvrDΔ40C) used in studies proposing the monomer helicase model. The study demonstrated that multiple UvrDΔ40C molecules jointly participated in DNA unwinding, presumably by forming an oligomer. Thus, the single-molecule observation addressed how the C-terminal amino acids affect the number of helicases bound to DNA, oligomerization, and unwinding activity, which can be applied to other helicases.

scholarly journals Regulation of Expression of the TIR-Containing Protein C Gene of the Uropathogenic Escherichia coli Strain CFT073

Pathogens ◽  
2021 ◽  
Vol 10 (5) ◽  
pp. 549
Author(s):  
Julia Ittensohn ◽  
Jacqueline Hemberger ◽  
Hannah Griffiths ◽  
Maren Keller ◽  
Simone Albrecht ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Protein C ◽  
Interleukin 1 ◽  
Coli Strain ◽  
Uropathogenic Escherichia Coli ◽  
Reporter Construct ◽  
Regulation Of Expression ◽  
E Coli ◽  
Strain Cft073 ◽  
Myeloid Differentiation Factor

The uropathogenic Escherichia coli strain CFT073 causes kidney abscesses in mice Toll/interleukin-1 receptor domain-containing protein C (TcpC) dependently and the corresponding gene is present in around 40% of E. coli isolates of pyelonephritis patients. It impairs the Toll-like receptor (TLR) signaling chain and the NACHT leucin-rich repeat PYD protein 3 inflammasome (NLRP3) by binding to TLR4 and myeloid differentiation factor 88 as well as to NLRP3 and caspase-1, respectively. Overexpression of the tcpC gene stopped replication of CFT073. Overexpression of several tcpC-truncation constructs revealed a transmembrane region, while its TIR domain induced filamentous bacteria. Based on these observations, we hypothesized that tcpC expression is presumably tightly controlled. We tested two putative promoters designated P1 and P2 located at 5′ of the gene c2397 and 5′ of the tcpC gene (c2398), respectively, which may form an operon. High pH and increasing glucose concentrations stimulated a P2 reporter construct that was considerably stronger than a P1 reporter construct, while increasing FeSO4 concentrations suppressed their activity. Human urine activated P2, demonstrating that tcpC might be induced in the urinary tract of infected patients. We conclude that P2, consisting of a 240 bp region 5′ of the tcpC gene, represents the major regulator of tcpC expression.

scholarly journals Structural genes for nitrate-inducible formate dehydrogenase in Escherichia coli K-12.

Genetics ◽  
1990 ◽  
Vol 125 (4) ◽  
pp. 691-702 ◽  
Author(s):  
B L Berg ◽  
V Stewart
Keyword(s):  
Escherichia Coli ◽  
Nitrate Reductase ◽  
Formate Dehydrogenase ◽  
Recombinant Dna ◽  
Structural Genes ◽  
Respiratory Pathway ◽  
E Coli ◽  
Formate Oxidation ◽  
Operon Expression ◽  
K 12

Abstract Formate oxidation coupled to nitrate reduction constitutes a major anaerobic respiratory pathway in Escherichia coli. This respiratory chain consists of formate dehydrogenase-N, quinone, and nitrate reductase. We have isolated a recombinant DNA clone that likely contains the structural genes, fdnGHI, for the three subunits of formate dehydrogenase-N. The fdnGHI clone produced proteins of 110, 32 and 20 kDa which correspond to the subunit sizes of purified formate dehydrogenase-N. Our analysis indicates that fdnGHI is organized as an operon. We mapped the fdn operon to 32 min on the E. coli genetic map, close to the genes for cryptic nitrate reductase (encoded by the narZ operon). Expression of phi(fdnG-lacZ) operon fusions was induced by anaerobiosis and nitrate. This induction required fnr+ and narL+, two regulatory genes whose products are also required for the anaerobic, nitrate-inducible activation of the nitrate reductase structural gene operon, narGHJI. We conclude that regulation of fdnGHI and narGHJI expression is mediated through common pathways.

scholarly journals Distribution of Lactoferrin Is Related with Dynamics of Neutrophils in Bacterial Infected Mice Intestine

Molecules ◽  
2020 ◽  
Vol 25 (7) ◽  
pp. 1496 ◽  
Author(s):  
Li Liang ◽  
Zhen-Jie Wang ◽  
Guang Ye ◽  
Xue-You Tang ◽  
Yuan-Yuan Zhang ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Antimicrobial Activity ◽  
Mucosal Immunity ◽  
Mrna Levels ◽  
Iron Binding ◽  
E Coli ◽  
New Knowledge ◽  
Activated Neutrophils ◽  
Binding Glycoprotein

Lactoferrin (Lf) is a conserved iron-binding glycoprotein with antimicrobial activity, which is present in secretions that recover mucosal sites regarded as portals of invaded pathogens. Although numerous studies have focused on exogenous Lf, little is known about its expression of endogenous Lf upon bacterial infection. In this study, we investigated the distribution of Lf in mice intestine during Escherichia coli (E. coli) K88 infection. PCR and immunohistology staining showed that mRNA levels of Lf significantly increased in duodenum, ileum and colon, but extremely decreased in jejunum at 8 h and 24 h after infection. Meanwhile, endogenous Lf was mostly located in the lamina propria of intestine villi, while Lf receptor (LfR) was in the crypts. It suggested that endogenous Lf-LfR interaction might not be implicated in the antibacterial process. In addition, it was interesting to find that the infiltration of neutrophils into intestine tissues was changed similarly to Lf expression. It indicated that the variations of Lf expression were rather due to an equilibrium between the recruitment of neutrophils and degranulation of activated neutrophils. Thus, this new knowledge will pave the way to a more effective understanding of the role of Lf in intestinal mucosal immunity.

SATP (YaaH), a succinate–acetate transporter protein in Escherichia coli

2013 ◽  
Vol 454 (3) ◽  
pp. 585-595 ◽  
Author(s):  
Joana Sá-Pessoa ◽  
Sandra Paiva ◽  
David Ribas ◽  
Inês Jesus Silva ◽  
Sandra Cristina Viegas ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Acetic Acid ◽  
Succinic Acid ◽  
Critical Role ◽  
Amino Acid Residues ◽  
E Coli ◽  
Transporter Protein ◽  
Affinity Constants ◽  
In Trans

In the present paper we describe a new carboxylic acid transporter in Escherichia coli encoded by the gene yaaH. In contrast to what had been described for other YaaH family members, the E. coli transporter is highly specific for acetic acid (a monocarboxylate) and for succinic acid (a dicarboxylate), with affinity constants at pH 6.0 of 1.24±0.13 mM for acetic acid and 1.18±0.10 mM for succinic acid. In glucose-grown cells the ΔyaaH mutant is compromised for the uptake of both labelled acetic and succinic acids. YaaH, together with ActP, described previously as an acetate transporter, affect the use of acetic acid as sole carbon and energy source. Both genes have to be deleted simultaneously to abolish acetate transport. The uptake of acetate and succinate was restored when yaaH was expressed in trans in ΔyaaH ΔactP cells. We also demonstrate the critical role of YaaH amino acid residues Leu131 and Ala164 on the enhanced ability to transport lactate. Owing to its functional role in acetate and succinate uptake we propose its assignment as SatP: the Succinate–Acetate Transporter Protein.

scholarly journals Molecular survey of mcr1 and mcr2 plasmid mediated colistin resistance genes in Escherichia coli isolates of animal origin in Iran

2021 ◽  
Vol 14 (1) ◽  
Author(s):  
Kayhan Ilbeigi ◽  
Mahdi Askari Badouei ◽  
Hossein Vaezi ◽  
Hassan Zaheri ◽  
Sina Aghasharif ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Resistance Genes ◽  
Companion Animals ◽  
Multidrug Resistant ◽  
Animal Origin ◽  
Colistin Resistance ◽  
E Coli ◽  
High Level ◽  
Farming Industry

Abstract Objectives The emergence of colistin-resistant Enterobacteriaceae from human and animal sources is one of the major public health concerns as colistin is the last-resort antibiotic for treating infections caused by multidrug-resistant Gram-negative bacteria. We aimed to determine the prevalence of the prototype widespread colistin resistance genes (mcr-1 and mcr-2) among commensal and pathogenic Escherichia coli strains isolated from food-producing and companion animals in Iran. Results A total of 607 E. coli isolates which were previously collected from different animal sources between 2008 and 2016 used to uncover the possible presence of plasmid-mediated colistin resistance genes (mcr-1 and mcr-2) by PCR. Overall, our results could not confirm the presence of any mcr-1 or mcr-2 positive E. coli among the studied isolates. It is concluded that despite the important role of food-producing animals in transferring the antibiotic resistance, they were not the main source for carriage of mcr-1 and mcr-2 in Iran until 2016. This study suggests that the other mcr variants (mcr-3 to mcr-9) might be responsible for conferring colistin resistance in animal isolates in Iran. The possible linkage between pig farming industry and high level of mcr carriage in some countries needs to be clarified in future prospective studies.

Sign in / Sign up

Export Citation Format

Share Document