Search PubMed⌕ Search

Biomedical subjects

R E Webster

Publications and source records attributed to R E Webster.

At least 19 recordsLinked to original sources
PubMed1992

Localization of TraC, a protein involved in assembly of the F conjugative pilus.

TraC is one of the proteins encoded by the F transfer region of the F conjugative plasmid which is required for the assembly of F pilin into the mature F pilus structure. Overproduction of this protein from the plasmid pKAS2, which carries only traC, resulted in the formation of inclusion bodies from which soluble TraC was purified. When small amounts of TraC were produced from pKAS2, the protein was localized to the cytoplasm by using anti-TraC antibodies. Similar analysis of a set of TraC-alkaline phosphatase fusion proteins localized all of these fusion proteins to the cytoplasm. However, when TraC was expressed from the F plasmid, much of it appeared associated with the bacterial membrane fraction. Under these conditions, TraC does not appear to be part of the tip of the F pilus, as neither anti-TraC antibodies nor purified TraC had any effect on the infection of F-containing bacteria by the filamentous bacteriophage f1. These data suggest that TraC is normally associated with the membrane through interactions with other proteins specified by the tra region. This interaction may be via the carboxyl-terminal region of the TraC protein, as a mutant TraC protein containing an Arg-Cys substitution at amino acid 811 exhibits an interaction with the membrane weaker than that of the wild-type protein in the presence of the other Tra proteins.

Alkaline Phosphatase↗
PubMed1992

Membrane localization and topology of a viral assembly protein.

The gene I protein (pI) of the filamentous bacteriophage f1 is required for the assembly of this virus. Antibodies specific to either the amino or carboxyl terminus of this protein were used to determine the location and topology of the gene I protein in f1-infected bacteria. pI is anchored in the inner membrane of Escherichia coli cells via a 20-amino-acid hydrophobic stretch, with its carboxyl-terminal 75 residues located in the periplasm and its amino-terminal 253 amino acids residing in the cytoplasm. By using the carboxyl-terminal pI antibody, a smaller protein, pI*, is also detected in f1-infected cells at a ratio of one to two molecules per molecule of pI. Analysis of proteins produced from a gene I amber mutant plasmid or bacteriophage suggests that pI* is most likely the result of an in-frame internal translational initiation event at methionine 241 of the 348-amino-acid pI. pI* is shown to be an integral inner membrane protein inserted in the same orientation as pI. The relation of the cellular locations of pI and pI* to some of the proposed functions of pI is discussed.

Base Sequence↗
PubMed1991

TolA: a membrane protein involved in colicin uptake contains an extended helical region.

The group A colicins and the DNA of many single-stranded filamentous bacteriophage are able to use combinations of the Tol proteins to gain entrance into or across the membrane of Escherichia coli. The TolA protein is a 421-amino acid residue integral membrane protein composed of three domains. Domain I, consisting of the amino-terminal 47 amino acids, contains a 21-residue hydrophobic segment that anchors the protein in the inner membrane. The remaining 374 amino acids, containing the other two domains, reside in the periplasmic space. Domain III, consisting of the carboxyl-terminal 120 residues, is considered to be the functional domain based on the location of the tolA592 deletion mutation. The internal 262 amino acids comprise domain II, which connects domains I and III together via short regions of polyglycine. It contains a large number of 3- to 5-residue polyalanine stretches, many of which have a repeat of the sequence Lys-Ala-Ala-Ala-(Glu/Asp). Circular dichroism analysis of different portions of TolA show domain II to be predominantly alpha-helical in structure while domain III contains approximately 10% helical structure.

Amino Acid Sequence↗
PubMed1991

The tol gene products and the import of macromolecules into Escherichia coli.

Genetic studies have identified a number of genes whose products appear to be required for the transport of the group A colicins and the single-stranded DNA of certain filamentous bacteriophages into Escherichia coli. Mutations in these genes allow normal binding of the colicins to their outer-membrane receptors and of the bacteriophage of the tip of specific conjugative pili, but do not allow translocation of the macromolecules to their target. These mutations have been designed 'tolerant' (tol) mutations and the protein products specified by these genes appear to comprise part of a transport system known as the Tol import system. Some of these genes have been isolated, sequenced and their protein products localized to the membranes or periplasm of E. coli. Information is also available regarding the domains of the colicins or phage proteins which interact with the Tol proteins. A preliminary model of the location and possible interactions of the Tol proteins is presented.

Bacterial Outer Membrane Proteins↗
PubMed1990

Nucleotide sequence of the F plasmid gene, traC, and identification of its product.

The traC gene of the F plasmid tra operon is required for the assembly of mature F-pilin subunits into extended F pili. The nucleotide sequence of traC was determined with a determined with a deduced coding region of 875 amino acids (aa) and 99066 Da. The traC1044 mutant allele, which allows filamentous phage infection in the absence of piliation, contains a C-to-T transition leading to an Arg----Cys substitution. Confirmation of the translational start came from the direct N-terminal aa sequencing of a TraC-alkaline phosphatase fusion protein.

Amino Acid Sequence↗
PubMed1989

Intravenous labetalol versus sodium nitroprusside for treatment of hypertension postcoronary bypass surgery.

Hypertension is common following coronary artery bypass surgery. The safety of labetalol, a recently released combined alpha-1 and beta-adrenergic blocking agent for treatment of hypertension in this clinical situation is controversial. The authors compared the hemodynamic effects of labetalol with those of sodium nitroprusside (SNP) in 91 patients with good left ventricular function and equally severe coronary artery disease and in whom coronary artery bypass surgery had been just completed. They were anesthetized using fentanyl, diazepam, and enflurane. If hypertension developed postoperatively, patients were randomized to receive labetalol, 2 mg/min to a maximum of 300 mg (20 patients) or sodium nitroprusside in 0.5 micrograms.kg-1.min-1 increments by infusion (20 patients) to return blood pressure to normal. Compared with control values, labetalol brought about significant (P less than 0.05) reductions in heart rate, and cardiac index. No change was noted in stroke volume or systemic vascular resistance, but slight increases were found in central venous pressure and pulmonary capillary wedge pressure. Sodium nitroprusside treatment caused significant increases in heart rate and cardiac index while reducing diastolic blood pressure, central venous pressure, and pulmonary capillary wedge pressure. Stroke volume remained unchanged. Following the study period, blood pressure was controlled in all patients with SNP. Total doses of SNP in the 16 h following the study period were significantly less in the labetalol group (46.6 +/- 11.7 mg) versus (116.1 +/- 10.3 mg) in the SNP group (P less than 0.05). In this clinical circumstance, labetalol can be safe and effective for controlling hypertension, but its mechanism of achieving this effect varies from that for sodium nitroprusside.(ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗
PubMed1989

Nucleotide sequences of the tolA and tolB genes and localization of their products, components of a multistep translocation system in Escherichia coli.

Various mutations in the tolQRAB gene cluster of Escherichia coli render the bacteria tolerant to high concentrations of the E, A, or K colicins as well as tolerant to infection by the single-stranded filamentous bacteriophage. The nucleotide sequence of a 2.8-kilobase fragment containing the tolA and tolB genes was determined. This sequence predicts TolA to be a 421-amino-acid protein of molecular mass 44,190 daltons. Studies using minicells show it to be associated with the inner membrane, presumably via a 21-amino-acid hydrophobic sequence between residues 13 and 35. The remaining 387 residues on the carboxyl side of this region are located in the periplasm. Within this region of TolA is a 230-residue portion that is predicted to form a very long helical segment. This region is rich in alanine, lysine, and glutamic and aspartic acids. The TolB protein is predicted to contain 431 amino acids. Localization studies using minicells show two proteins encoded by this open reading frame. The larger protein of 47.5 kilodaltons appears to be associated with the membrane fractions. The smaller protein is 43 kilodaltons in size and is found with the periplasmic components of the cell.

Amino Acid Sequence↗
PubMed1989

Leisure: how to promote inpatient motivation after discharge.

1. A group of psychiatric inpatients followed plans to participate in leisure activities after discharge at significantly higher levels when treated with classes and written contracts; or classes, written contracts, and musical entertainment; rather than with classes alone. 2. Inpatient classes for psychiatric patients should be coupled with other interventions, such as contracts or musical entertainment, to assist patients to attain therapeutic goals after discharge. 3. The utilization of written goal-setting contracts seems to have merit as a routine nursing intervention with psychiatric inpatients. 4. The use of a written contract left with the patient may promote or enhance a nurse/patient transaction.

Female↗
PubMed1988

An amino acid sequence which directs membrane insertion causes loss of membrane potential.

A 55-amino acid segment, normally present between residues 241 and 295 of the 348-residue gene I protein of the filamentous bacteriophage f1, acts as an internal signal sequence for gene I protein or, when present in fusion proteins, for EcoRI endonuclease or alkaline phosphatase. The resulting proteins are inserted so that they span the membrane with sequences on the amino side of the 55-residue segment in the cytoplasm and those near the carboxy side outside the cytoplasmic membrane. The presence of these proteins in the membrane results in the rapid inhibition of cell growth, probably from a loss of the membrane potential. We describe some of the elements in this 55-residue segment that appear to be crucial for its interaction with the membrane.

Amino Acid Sequence↗
PubMed1988

Low-frequency infection of F- bacteria by transducing particles of filamentous bacteriophages.

Filamentous particles containing single-stranded plasmid and bacteriophage DNA are able to infect F- Escherichia coli at frequencies of approximately 10(-6). This infection is dependent on an intact particle and requires the products of the tolQ, tolR, and tolA genes of the bacteria. The addition of CaCl2 can increase the frequency about 100-fold, presumably by increasing the concentration of particles at the bacterial surface.

Coliphages↗
PubMed1987

Nucleotide sequence of a gene cluster involved in entry of E colicins and single-stranded DNA of infecting filamentous bacteriophages into Escherichia coli.

Mutations in fii or tolA of the fii-tolA-tolB gene cluster at 17 min on the Escherichia coli map render cells tolerant to high concentrations of the E colicins and do not allow the DNA of infecting single-stranded filamentous bacteriophages to enter the bacterial cytoplasm. The nucleotide sequence of a 1,854-base-pair DNA fragment carrying the fii region was determined. This sequence predicts three open reading frames sequentially coding for proteins of 134, 230, and 142 amino acids, followed by the potential start of the tolA gene. Oligonucleotide mutagenesis of each open reading frame and maxicell analysis demonstrated that all open reading frames are expressed in vivo. Sequence analysis of mutant fii genes identified the 230-amino acid protein as the fii gene product. Chromosomal insertion mutations were constructed in each of the two remaining open reading frames. The phenotype resulting from an insertion of the chloramphenicol gene into the gene coding for the 142-amino acid protein is identical to that of mutations in fii and tolA. This gene is located between fii and tolA, and we propose the designation of tolQRA for this cluster in which tolQ is the former fii gene and tolR is the new open reading frame. The protein products of this gene cluster play an important role in the transport of large molecules such as the E colicins and filamentous phage DNA into the bacterium.

Amino Acid Sequence↗
PubMed1987

A traC mutant that retains sensitivity to f1 bacteriophage but lacks F pili.

An F lac pro mutant which was temperature sensitive for infection by the filamentous bacteriophage f1 but resistant to the F-specific icosahedral RNA phage f2 was isolated. Cells carrying the F' mutation failed to elaborate F pili at all temperatures. Mutant cells were able to pair with recipient cells during bacterial conjugation, but transfer of conjugal DNA occurred at a greatly reduced frequency. Complementation analyses showed the F' mutation to be in the traC gene. When a plasmid carrying traC was introduced into hosts harboring the F' mutation, phage sensitivity, the ability to elaborate F pili, and conjugation efficiency were restored. The mutation was named traC1044. The F lac pro traC1044 mutant appears to be unique among traC mutants in retaining host sensitivity to the filamentous phage f1 in the absence of expression of extended F pili. Phage f1 attachment sites appeared to be present at the cell surface in traC1044 mutants. The reduced accessibility of these sites may account for the reduced efficiency of phage f1 infection of traC1044 hosts, although the possibility that a defect was present in the receptor site itself was not eliminated. Membranes of hosts carrying the F' mutation contained a full complement of mature F-pilin subunits, so the product of traC is presumably required for pilus assembly but not for pilin processing. This, together with the deficiency in conjugal DNA transfer, suggests that traC may be part of a membrane-spanning tra protein complex responsible for pilus assembly and disassembly and conjugal DNA transmission.

Chromosome Mapping↗
PubMed1986

Morphogenesis of f1 filamentous bacteriophage. Increased expression of gene I inhibits bacterial growth.

We have cloned the gene I sequence of the filamentous bacteriophage f1 downstream from the lambda leftward promoter on a plasmid that also contains the temperature-sensitive lambda repressor, cI857. Temperature induction of gene I protein (pI) resulted in rapid cessation of growth. This inhibition appears to involve a rapid decrease in synthesis of host protein and RNA. The ability of pI to cause this inhibition is not dependent on thioredoxin, a host factor that is necessary for phage morphogenesis and has been shown by genetic data to interact with pI. The inhibition does not appear to be mediated by the amino half of the protein, as induction of an identical plasmid construction of an amber mutant positioned two-thirds along gene I, does not affect cell growth. Analysis of the transcription products from the cloned gene I confirmed previous suggestions that a transcription terminator exists in the amino-terminal portion of the gene. In addition, there is no detectable promoter activity in the 152 bases immediately upstream from the gene. These data and the inability to overproduce pI argue for down-regulation of pI production. Radioactive labeling of proteins in maxi-cells and normal Escherichia coli cells identifies pI as a protein of about 39,000 Mr that partitions with the cell envelope. Pulse-chase experiments suggest that pI is not processed to any appreciable extent.

Bacteriophages↗
PubMed1986

fii, a bacterial locus required for filamentous phage infection and its relation to colicin-tolerant tolA and tolB.

We describe mutations in a new bacterial locus, designated fii, which do not allow the filamentous bacteriophage f1 to infect bacteria harboring the F plasmid. Mutations at this locus do not affect the ability of F plasmid-containing bacteria to undergo conjugation or be infected by the F plasmid-specific RNA phage f2. The filamentous phage can still adsorb to the F sex pilus, but the DNA is unable to enter the bacteria. All fii mutants become tolerant to colicins E1, E2, and E3. Strains with amber mutations in fii also are unable to plaque P1, even though they can be infected with this phage. Mutations in fii also prevent infection of bacteria harboring the N plasmid by the filamentous bacteriophage IKe. The fii locus maps adjacent to tolA, mutants of which demonstrate tolerance to high levels of the E and K colicins. The three genes tolA, tolB, and fii are shown to reside on a 4.3-kilobase fragment of the Escherichia coli chromosome. Each gene has been cloned into a chimeric plasmid and shown to complement, in trans, mutations at the corresponding chromosomal locus. Studies in maxicells show that the product of fii appears to be a 24-kilodalton protein which copurifies with the cell envelope. The product of tolA has been identified tentatively as a 51-kilodalton protein. Data from cloning, Tn5 mutagenesis, and P1 transduction studies are consistent with the gene order sucA-fii-tolA-tolB-aroG near 17 min on the E. coli map.

Bacterial Proteins↗
PubMed1985

An in-vitro investigation of synergy and antagonism between antimicrobials against Chlamydia trachomatis.

Chequerboard titrations of antimicrobials were carried out against Chlamydia trachomatis in vitro to assess possible synergy or antagonism. None of the antimicrobial pairs produced any detectable antagonism. Penicillin and ciprofloxacin showed independent activity. Tetracycline and penicillin, tetracycline and erythromycin, tetracycline and chloramphenicol, erythromycin and penicillin, and erythromycin and chloramphenicol showed additive inhibitory activity and limited synergy. Trimethoprim and sulphamethoxazole was the only combination to produce clear synergistic activity against Chlam. trachomatis in vitro.

Anti-Bacterial Agents↗