SEARCH · PubMed Health

Results for “Operon”

Explore indexed PubMed citations for clinical trials, systematic reviews and public health research. Read source abstracts and follow each citation to its original PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 1,477 records · Page 82Linked to original sources

PubMed1991

Role and expression of the Bacillus subtilis rodC operon.

The role of the rodC operon in Bacillus subtilis was investigated. The operon encodes two genes (rodD and rodC) necessary for the synthesis of the cell wall teichoic acid. Transcription of this operon is responsive to levels of phosphate and to concentrations of magnesium ions in the growth medium. This regulation of mRNA production corresponds to conditions that dictate the type of polymer that will be synthesized for the cell wall, i.e., teichoic or teichuronic acid. While the introduction of multiple copies of rodC was tolerated by the cells, multiple copies of rodD appeared to be lethal. The lethality of the rodD fragment was not exhibited if multiple copies of rodC were also present.

Bacillus subtilis↗

PubMed1991

Anaerobic regulation of transcription initiation in the arcDABC operon of Pseudomonas aeruginosa.

The arcDABC operon of Pseudomonas aeruginosa encodes the enzymes of the arginine deiminase pathway, which is inducible under conditions of oxygen limitation and serves to generate ATP from arginine. The 5' end of arc mRNA extracted from anaerobically grown cells was determined by S1 and primer extension mapping. The transcription initiation site was located upstream of the arcD gene and 41.5 bp downstream of the center of the sequence TTGAC....ATCAG. This sequence, termed the ANR box, is similar to the consensus FNR recognition site of Escherichia coli. Transcription of the arc operon in P. aeruginosa was strongly decreased by a deletion of the TTGAC half site or by a mutation in the anr gene, which is known to code for the FNR-like regulatory protein ANR. During a transition from aerobic to anaerobic growth conditions, the concentrations of arc mRNAs and the levels of the ArcD and ArcA proteins rose in a parallel fashion. Mutational analysis of the arc promoter region led to the conclusion that the distance between the ANR box and the -10 promoter region is important for promoter strength, whereas the -35 region does not appear to be critical for arc promoter function. These findings and previous results indicate that anaerobic induction of the arc operon occurs at the level of transcription and requires the ANR box in cis and the ANR protein in trans.

Amino Acid Sequence↗

PubMed1991

Expression and operon structure of the sel genes of Escherichia coli and identification of a third selenium-containing formate dehydrogenase isoenzyme.

A detailed analysis of the expression of the sel genes, the products of which are necessary for the specific incorporation of selenium into macromolecules in Escherichia coli, showed that transcription was constitutive, being influenced neither by aerobiosis or anaerobiosis nor by the intracellular selenium concentration. The gene encoding the tRNA molecule which is specifically aminoacylated with selenocysteine (selC) proved to be monocistronic. In contrast, the other three sel genes (selA, -B, and -D) were shown to be constituents of two unlinked operons. The selA and selB genes formed one transcriptional unit (sel vector AB), while selD was shown to be the central gene in an operon including two other genes, the promoter distal of which (topB) encodes topoisomerase III. The promoter proximal gene (orf183) was sequenced and shown to encode a protein consisting of 183 amino acids (Mr, 20,059), the amino acid sequence of which revealed no similarity to any currently known protein. The products of the orf183 and topB genes were required neither for selenoprotein biosynthesis nor for selenation of tRNAs. selAB transcription was driven by a single, weak promoter; however, two major selD operon transcripts were identified. The longer initiated just upstream of the orf183 gene, whereas the 5' end of the other mapped in a 116-bp nontranslated region between orf183 and selD. Aerobic synthesis of all four sel gene products incited a reexamination of a weak 110-kDa selenopolypeptide which is produced under these conditions. The aerobic appearance of this 110-kDa selenopolypeptide was not a consequence of residual expression of the gene encoding the 110-kDa selenopolypeptide of the anaerobically inducible formate dehydrogenase N (FDHN) enzyme, as previously surmised, but rather resulted from the expression of a gene encoding a third, distinct selenopolypeptide in E. coli. A mutant strain no longer capable of synthesizing the 80- and 110-kDa selenopolypeptides of FDHH and FDHN, respectively, still synthesized this alternative 110-kDa selenopolypeptide which was present at equivalent levels in cells grown aerobically and anaerobically with nitrate. Furthermore, this strain exhibited a formate- and sel gene-dependent respiratory activity, indicating that it is probable that this selenopolypeptide constitutes a major component of the formate oxidase, an enzyme activity initially discovered in aerobically grown E. coli more than 30 years ago.

Amino Acid Sequence↗

PubMed1991

Multiple mechanisms contribute to osmotic inducibility of proU operon expression in Escherichia coli: demonstration of two osmoresponsive promoters and of a negative regulatory element within the first structural gene.

Transcription of the proU operon in Escherichia coli is induced several hundredfold upon growth of cells in media of elevated osmolarity. A low-copy-number promoter-cloning plasmid vector, with lacZ as the reporter gene, was used for assaying the osmoresponsive promoter activity of each of various lengths of proU DNA, generated by cloning of discrete restriction fragments and by an exonuclease III-mediated deletion approach. The results indicate that expression of proU in E. coli is directed from two promoters, one (P2) characterized earlier by other workers with the start site of transcription 60 nucleotides upstream of the initiation codon of the first structural gene (proV), and the other (P1) situated 250 nucleotides upstream of proV. Furthermore, a region of DNA within proV was shown to be involved in negative regulation of proU transcription; phage Mu dII1681-generated lac fusions in the early region of proV also exhibited partial derepression of proU regulation, in comparison with fusions further downstream in the operon. Sequences around promoter P1, sequences around P2, and the promoter-downstream negative regulatory element, respectively, conferred approximately 5-, 8-, and 25-fold osmoresponsivity on proU expression. Within the region genetically defined to encode the negative regulatory element, there is a 116-nucleotide stretch that is absolutely conserved between the proU operons of E. coli and Salmonella typhimurium and has the capability of exhibiting alternative secondary structure. Insertion of this region of DNA into each of two different plasmid vectors was associated with a marked reduction in the mean topological linking number in plasmid molecules isolated from cultures grown in high-osmolarity medium. We propose that this region of DNA undergoes reversible transition to an underwound DNA conformation under high-osmolarity growth conditions and that this transition mediates its regulatory effect on proU expression.

Base Sequence↗

PubMed1991

srfA is an operon required for surfactin production, competence development, and efficient sporulation in Bacillus subtilis.

The srfA locus of Bacillus subtilis is defined by a transposon Tn917 insertion and is required for production of the peptide secondary metabolite surfactin. The srfA locus was isolated by cloning the DNA flanking srfA::Tn917 insertions followed by chromosome walking. The cloned region is an operon of over 25 kb which covers the transcription initiation region but not the intact 3' end of srfA. csh-293, which was previously identified as a Tn917lac mutation that impairs competence development and causes a conditional defect in sporulation, was known to be located in the vicinity of the srfA locus within the B. subtilis genome. The csh-293::Tn917lac mutation was discovered to cause a defect in surfactin production and was shown to be located in the srfA operon by its cotransformation with srfA mutations and by Southern hybridization analysis. Insertion mutations in srfA, created by the chromosomal integration of plasmids bearing overlapping srfA DNA fragments, were examined for their effects on surfactin production, competence, and sporulation. All three processes were found to require the intact 5' half of the srfA operon, whereas the 3' half of srfA was found to be required for sporulation and surfactin production but not competence. These experiments show that srfA gene products function in B. subtilis cell specialization and differentiation.

Amino Acid Sequence↗

PubMed1991

Physical map of the chromosome of Lactococcus lactis subsp. lactis DL11 and localization of six putative rRNA operons.

A physical map of the chromosome of Lactococcus lactis subsp. lactis DL11 was constructed by using the contour-clamped homogeneous electric field mode of pulsed-field gel electrophoresis in one- and two-dimensional separations to analyze restriction digests of high-molecular-weight genomic DNA. The map, which shows all the observed NotI and SmaI sites (six and 21, respectively) and 8 of approximately 30 SalI sites, is circular and yields a total size of 2.58 megabase pairs for the L. lactis subsp. lactis DL11 chromosome. By using rDNA from Mycoplasma capricolum to probe Southern blots of pulsed-and fixed-field digestion patterns, six putative rRNA operons were identified in L. lactis subsp. lactis DL11 and placed on the map of the chromosome. Five of these loci are clustered in a region representing only 20% of the chromosome. The presence of a SmaI site in each of the putative operons allowed the direction of transcription of each operon to be deduced.

Blotting, Southern↗

PubMed1992

Cloning, sequencing, and molecular analysis of the groESL operon of Clostridium acetobutylicum.

The groESL operon of Clostridium acetobutylicum was cloned in Escherichia coli by using a gene probe of E. coli groESL. Sequencing of a positively reacting 2.2-kbp HindIII fragment contained in the recombinant plasmid pFN1 and a 2.5-kbp XbaI fragment present in pFN4 revealed that both fragments partially overlapped and together spanned 3,493 bp of the clostridial chromosome. Two complete open reading frames (288 and 1632 bp) were found and identified as the groES- and groEL-homologous genes of C. acetobutylicum, respectively. The 3' end of a third gene (orfZ), which was divergently transcribed, showed no significant homology to other sequences available in the EMBL and GenBank data bases. The length of the groESL-specific mRNA (2.2 kb), a transcription terminator downstream of groEL, and a transcription start site upstream of groES, identified by primer extension analysis, indicated that groES and groEL of C. acetobutylicum are organized in a bicistronic operon. From the transcription start site, the promoter structure 5'-TTGCTA (17 bp) TATTAT that shows high homology to the consensus promoter sequence of gram-positive bacteria as well as E. coli was deduced. Transcription of the groESL operon was strongly heat inducible, and maximum levels of mRNA were detected 15 min after heat shock from 30 to 42 degrees C. An 11-bp inverted repeat, located between promoter and translation start sites of groES and partially identical with similar structures in front of several heat shock genes of other bacteria, may play an important role in the regulation of heat shock gene expression in this organism.

Amino Acid Sequence↗

PubMed1992

The Escherichia coli K-12 cyn operon is positively regulated by a member of the lysR family.

A regulatory gene, cynR, was found to be located next to the cyn operon but transcribed in the opposite direction. cynR encodes a positive regulatory protein that controls the cyn operon as well as its own synthesis. Positive regulation of the cyn operon requires cyanate and the cynR protein, but the negative autoregulation of the cynR gene appears to be independent of cyanate. The predicted amino acid sequence of the cynR protein derived from the DNA sequence was found to have significant homology to the predicted amino acid sequence of the lysR family of regulatory proteins.

Amino Acid Sequence↗

PubMed1992

Cloning and characterization of the groESL operon from Bacillus subtilis.

The sequence of the 10 N-terminal amino acids of a Bacillus subtilis protein that cross-reacts with antibody to Escherichia coli GroEL was used to design a set of degenerate oligonucleotide probes. These probes identified a clone which carries almost the entire groESL operon from a B. subtilis subgenomic library. By chromosomal walking, an additional fragment carrying the 3' end of groESL and its flanking sequence was isolated. Sequence analysis revealed two open reading frames (ORFs) in the cloned DNA. The upstream ORF encodes a 10-kDa protein which has 47% amino acid identity with E. coli GroES. The downstream ORF encodes a 58-kDa protein which is 62% identical to E. coli GroEL. A 2.1-kb groESL mRNA from B. subtilis was detected independently by Northern (RNA) blot analyses with a groES- and a groEL-specific probe. This demonstrated that groES and groEL are in an operon. The groESL promoter was located by using a promoter-probing plasmid, and the apparent transcription start site was mapped by primer extension analysis. The same promoter is utilized under normal and heat shock conditions. This promoter has the same features as a typical sigma A promoter. A strain in which the groESL operon was under the control of the sucrose-inducible sacB promoter was created. With this strain, it was possible to show that both groES and groEL are essential genes under both normal and heat shock conditions.

Amino Acid Sequence↗

PubMed1992

arc-dependent thermal regulation and extragenic suppression of the Escherichia coli cytochrome d operon.

In a screen for Escherichia coli genes whose products are required for high-temperature growth, we identified and characterized a mini-Tn10 insertion that allows the formation of wild-type-size colonies at 30 degrees C but results in microcolony formation at 36 degrees C and above (Ts- phenotype). Mapping, molecular cloning, and DNA sequencing analyses showed that the mini-Tn10 insertion resides in the cydB gene, the distal gene of the cydAB operon (cytochrome d). The Ts- growth phenotype was also shown to be associated with previously described cyd alleles. In addition, all cyd mutants were found to be extremely sensitive to hydrogen peroxide. Northern (RNA) blot analysis showed that cyd-specific mRNA levels accumulate following a shift to high temperature. Interestingly, this heat shock induction of the cyd operon was not affected in an rpoH delta background but was totally absent in an arcA or arcB mutant background. Extragenic suppressors of the Cyd Ts- phenotype are found at approximately 10(-3). Two extragenic suppressors were shown to be null alleles in either arcA or arcB. One interpretation of our results is that in the absence of ArcA or ArcB, which are required for the repression of the cyo operon (cytochrome o), elevated levels of Cyo are produced, thus compensating for the missing cytochrome d function. Consistent with this interpretation, the presence of the cyo gene on a multicopy plasmid suppressed the Ts- and hydrogen peroxide-sensitive phenotypes of cyd mutants.

Bacterial Outer Membrane Proteins↗

PubMed1992

Analysis of operons encoding 23S rRNA of Clostridium botulinum type A.

Southern hybridization analysis of Clostridium botulinum type A chromosomal DNA indicated the presence of six copies of the 23S rRNA gene. Fragments of DNA encoding 23S rRNA were amplified by polymerase chain reaction and cloned in Escherichia coli. Three clones examined by restriction enzyme and sequence analysis were found to be derived from different operons. Sequence determination of the entire insert of two clones revealed nine nucleotide changes in the genes coding for 23S rRNA (99.7% sequence identity) between operons encoded on the same chromosome, showing microheterogeneity in the rRNA operons of this organism.

Base Sequence↗

PubMed1992

Suppression of insertions in the complex pdxJ operon of Escherichia coli K-12 by lon and other mutations.

Complementation analyses using minimal recombinant clones showed that all known pdx point mutations, which cause pyridoxine (vitamin B6) or pyridoxal auxotrophy, are located in the pdxA, pdxB, serC, pdxJ, and pdxH genes. Antibiotic enrichments for chromosomal transposon mutants that require pyridoxine (vitamin B6) or pyridoxal led to the isolation of insertions in pdxA, pdxB, and pdxH but not in pdxJ. This observation suggested that pdxJ, like pdxA, pdxB, and serC, might be in a complex operon. To test this hypothesis, we constructed stable insertion mutations in and around pdxJ in plasmids and forced them into the bacterial chromosome. Physiological properties of the resulting insertion mutants were characterized, and the DNA sequence of pdxJ and adjacent regions was determined. These combined approaches led to the following conclusions: (i) pdxJ is the first gene in a two-gene operon that contains a gene, temporarily designated dpj, essential for Escherichia coli growth; (ii) expression of the rnc-era-recO and pdxJ-dpj operons can occur independently, although the pdxJ-dpj promoter may lie within recO; (iii) pdxJ encodes a 26,384-Da polypeptide whose coding region is preceded by a PDX box, and dpj probably encodes a basic, 14,052-Da polypeptide; (iv) mini-Mud insertions in dpj and pdxJ, which are polar on dpj, severely limit E. coli growth; and (v) three classes of suppressors, including mutations in lon and suppressors of lon, that allow faster growth of pdxJ::mini-Mud mutants can be isolated. A model to account for the action of dpj suppressors is presented, and aspects of this genetic analysis are related to the pyridoxal 5'-phosphate biosynthetic pathway.

ATP-Dependent Proteases↗

PubMed1993

Regulation of kdp operon expression in Escherichia coli: evidence against turgor as signal for transcriptional control.

Kdp, an inducible high-affinity K+ transporter in Escherichia coli, is encoded by genes of the kdpABC operon, and its expression is regulated by the products of kdpD and kdpE. Loss of cell turgor has been proposed to be the signal which induces kdp expression (L. A. Laimins, D. B. Rhoads, and W. Epstein, Proc. Natl. Acad. Sci. USA 78:464-468, 1981). We reexamined kdp expression during steady-state growth under a variety of conditions and were able to confirm earlier observations which had indicated that it is primarily affected by the concentration of K+ in the medium and by mutations in genes encoding various K+ transporters in E. coli. Changes in pH of the culture also altered kdp expression. In all of these cases, an increase in [K+] of the medium repressed the operon. Several ionic solutes induced steady-state kdp expression (but to differing extents), whereas nonionic solutes had no effect, indicating that kdp expression is not determined by osmolarity of the growth medium. kdp expression during steady-state growth was shown also to be unaffected by the accumulation of other intracellular compatible solutes such as trehalose or glycine betaine, which would be expected to restore cell turgor during growth in high-osmolarity media. Two mutants that are defective in perception of the signal regulating kdp were isolated, and the mutation in each of them was mapped to the kdpDE regulatory locus. Analysis of kdp expression in one of these mutants provided additional evidence against the turgor regulation model. On the basis of these data, we discuss alternative candidates that might serve as the signal for control of kdp operon transcription.

Adenosine Triphosphatases↗

PubMed1993

Mutations that relieve nutritional repression of the Bacillus subtilis dipeptide permease operon.

The Bacillus subtilis dciA operon encodes a dipeptide transport complex that is induced rapidly as cells enter stationary phase and initiate sporulation. Expression of this operon in growing cells is repressed by glucose, by a mixture of amino acids, and by the AbrB protein. A genetic screen was devised to identify mutations that allow inappropriate expression from the dciA promoter during growth. These mutations resulted in increased dciA transcription during growth in nutrient broth, in minimal amino acids medium, and in minimal glucose medium. Some of the mutations, called dcs (dciA control site), were cloned and shown by sequence analysis to cluster near the start site of dciA transcription. Primer extension and in vitro transcription analysis revealed that the dcs mutations did not create a new promoter. These mutations may therefore disrupt an operator site necessary for the binding of a negative regulator responsive to the nutritional state of the cell. The dcs mutant promoters were still subject to AbrB control, suggesting that the dciA operon is regulated by at least two proteins, AbrB and a nutritionally responsive regulator. The gene(s) for the putative nutritional regulator may be defined by the cod (control of dciA) mutations, which appeared to relieve amino acid and glucose repression of dciA by altering a diffusible factor. An abrB cod double mutant exhibited high-level expression of dciA during exponential growth phase.

Amino Acids↗

PubMed1993

The Bacillus subtilis ochre suppressor sup-3 is located in an operon of seven tRNA genes.

Most Bacillus subtilis tRNA genes have been isolated from lambda libraries by use of probes that hybridize to tRNA or rRNA sequences. None of those genes map to the region of the sup-3 mutation. By cloning of the sup-3 allele, a cluster of seven tRNA genes (the trnS operon) that had not been isolated by other methods was identified. In principle, this approach could be used to isolate at least one more predicted tRNA-containing operon in this bacterium. The trnS operon was shown to contain tRNA genes for Asn (GUU), Ser (GCU), Glu (UUC), Gln (UUG), Lys (UUU), Leu (UAG), and Leu (GAG). The sup-3 mutation was found to be a T-to-A transversion that changes the anticodon of the lysine tRNA from 5'-UUU-3' to 5'-UUA-3'. This result agrees with previous work that determined that the sup-3 mutation causes lysine to be inserted at ochre nonsense mutations.

Bacillus subtilis↗

PubMed1993

Use of the rep technique for allele replacement to construct mutants with deletions of the pstSCAB-phoU operon: evidence of a new role for the PhoU protein in the phosphate regulon.

The phosphate regulon is negatively regulated by the PstSCAB transporter and PhoU protein by a mechanism that may involve protein-protein interaction(s) between them and the Pi sensor protein, PhoR. In order to study such presumed interaction(s), mutants with defined deletions of the pstSCAB-phoU operon were made. This was done by construction of M13 recombinant phage carrying these mutations and by recombination of them onto the chromosome by using a rep host (which cannot replicate M13) for allele replacement. These mutants were used to show that delta (pstSCAB-phoU) and delta (pstB-phoU) mutations abolished Pi uptake by the PstSCAB transporter, as expected, and that delta phoU mutations had no effect on uptake. Unexpectedly, delta phoU mutations had a severe growth defect, and this growth defect was (largely) alleviated by a compensatory mutation in the pstSCAB genes or in the phoBR operon, whose gene products positively regulate expression of the pstSCAB-phoU operon. Because delta phoU mutants that synthesize a functional PstSCAB transporter constitutively grew extremely poorly, the PhoU protein must have a new role, in addition to its role as a negative regulator. A role for the PhoU protein in intracellular Pi metabolism is proposed. Further, our results contradict those of M. Muda, N. N. Rao, and A. Torriani (J. Bacteriol. 174:8057-8064, 1992), who reported that the PhoU protein was required for Pi uptake.

Alleles↗

PubMed1993

The cbb operons of the facultative chemoautotroph Alcaligenes eutrophus encode phosphoglycolate phosphatase.

The two highly homologous cbb operons of Alcaligenes eutrophus H16 that are located on the chromosome and on megaplasmid pHG1 contain genes encoding several enzymes of the Calvin carbon reduction cycle. Sequence analysis of a region from the promoter-distal part revealed two open reading frames, designated cbbT and cbbZ, at equivalent positions within the operons. Comparisons with known sequences suggested cbbT to encode transketolase (TK; EC 2.2.1.1) as an additional enzyme of the cycle. No significant overall sequence similarities were observed for cbbZ. Although both regions exhibited very high nucleotide identities, 93% (cbbZ) and 96% (cbbT), only the chromosomally encoded genes were heterologously expressed to high levels in Escherichia coli. The molecular masses of the observed gene products, CbbT (74 kDa) and CbbZ (24 kDa), correlated well with the values calculated on the basis of the sequence information. TK activities were strongly elevated in E. coli clones expressing cbbT, confirming the identity of the gene. Strains of E. coli harboring the chromosomal cbbZ gene showed high levels of activity of 2-phosphoglycolate phosphatase (PGP; EC 3.1.3.18), a key enzyme of glycolate metabolism in autotrophic organisms that is not present in wild-type E. coli. Derepression of the cbb operons during autotrophic growth resulted in considerably increased levels of TK activity and the appearance of PGP activity in A. eutrophus, although the pHG1-encoded cbbZ gene was apparently not expressed. To our knowledge, this study represents the first cloning and sequencing of a PGP gene from any organism.

Alcaligenes↗

PubMed1993

Salicylate induction of antibiotic resistance in Escherichia coli: activation of the mar operon and a mar-independent pathway.

Since the growth of wild-type Escherichia coli in salicylate results in a multiple antibiotic resistance phenotype similar to that of constitutive mutants (Mar) of the chromosomal mar locus, the effect of salicylate on the expression of the marRAB operon was investigated. The amount of RNA hybridizing with a mar-specific DNA probe was 5 to 10 times higher in wild-type cells grown with sodium salicylate (5.0 mM) than in untreated controls. Untreated Mar mutants had three to five times more mar-specific RNA than wild-type cells did. When a Mar mutant was treated with salicylate, a 30- to 50-fold increase of mar-specific RNA was seen. In wild-type cells bearing a mar promoter-lacZ fusion on the chromosome, salicylate increased beta-galactosidase activity by sixfold. Thus, salicylate induces transcription of the marRAB operon. Other inducers of phenotypic multiple antibiotic resistance, e.g., benzoate, salicyl alcohol, and acetaminophen, but not acetate, also increased transcription from the mar promoter but to a lesser extent than did salicylate. Both in wild-type and mar-deficient strains, growth in salicylate resulted in increased antibiotic resistance, decreased permeation of the outer membrane to cephaloridine, increased micF transcription, and decreased amounts of OmpF. However, the magnitude of these changes was generally greater in wild-type (mar-containing) cells. Thus, salicylate and other compounds can induce transcription of the mar operon and, presumably, give rise to multiple antibiotic resistance via this pathway. However, salicylate can also activate an unidentified, mar-independent pathway(s) which engenders multiple antibiotic resistance.

Ampicillin↗