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Biomedical subjects

J R Walker

Publications and source records attributed to J R Walker.

At least 127 records · Page 7Linked to original sources
PubMed1984

An E. coli DNA fragment 118 base pairs in length provides dnaY+ complementing activity.

The dnaY gene of E. coli, thought to be involved in the polymerization phase of DNA replication, was localized on a fragment 118 base pairs in length. This fragment, cloned in two different vectors and tested in a dnaY (Ts) recA host, has dnaY + complementing activity. The nucleotide sequence of the 118 base pairs and flanking bases was determined. The dnaY complementing activity was inactivated by transposon insertion and by localized chemical mutagenesis. Three independent insertions of Tn5 into the 118 base pair region eliminated dnaY activity. Eight single-base-change mutations that resulted in loss of dnaY activity also were located within the 118 base pair region. Analysis of the nucleotide sequence reveals a potential promoter but reveals no open reading frames likely to be translated into polypeptides. However, an RNA transcript of the dnaY region is synthesized in vivo. Perhaps the active product of dnaY is a small RNA or perhaps the dnaY region functions as a site.

Base Composition↗
PubMed1983

Cloning of the Escherichia coli dnaZX region and identification of its products.

The Escherichia coli DNA replication genes dnaZ and dnaX have previously been localized very near each other at 10.4 to 10.5 min on the chromosome map. These genes were cloned from a dnaZ+X+ plasmid of the Clarke and Carbon collection by identifying complementing fragments and both were located on a 2.1 kilobase pair (kb) fragment. The organization of the Z and X genes was investigated by Tn5 mutagenesis of a Z+X+ plasmid. Insertions which abolished Z or X complementing activity were mapped by restriction enzyme analysis within the 2.1 kb fragment. With the exception of one atypical insertion, all the insertions inactivated both Z and X complementation. The protein products of the dnaZ-dnaX region were labelled in minicells containing dnaZ+X+ and dnaZX::Tn5 plasmids. The 2.1 kb ZX region (which has a maximum coding capacity of 77,000 daltons of protein in a single reading frame) directed the synthesis of two proteins, one of 75,000 daltons, designated dnaX, and another of 56,500 daltons, designated dnaZ. Tn5 insertion into the ZX region interrupted the synthesis of these proteins; the detection of truncated fragments of dnaX determined the direction of transcription. In vitro, using a coupled transcription-translation system dependent on plasmid DNA, synthesis of the 75,000 dalton dnaX protein was demonstrated, but there was no detectable synthesis of the smaller dnaZ protein. Probably, therefore, the 75,000 dalton dnaX protein is cleaved in vivo to generate the dnaZ protein. It is possible that the 75,000 dalton product is the tau subunit of DNA polymerase III because they migrated similarly in electrophoresis.

Bacterial Proteins↗
PubMed1983

Reduced susceptibility to natural killer cell lysis of hamster tumours exhibiting high levels of spontaneous metastasis.

The natural killer (NK) cell susceptibility of hamster tumours exhibiting high and low levels of spontaneous metastasis in vivo was investigated. The parent cell line (HSV-2-333-2-26), which was weakly metastatic, and the NK-sensitive target line K562 were both sensitive to hamster natural killer cell activity. In contrast, 2 in vivo cloned sublines, designated met A and met B, exhibited a high level of spontaneous metastasis and were shown to be weakly or non-susceptible to hamster UK reactivity.

Animals↗
PubMed1983

Human lymphoblastoid interferon in the treatment of small cell lung cancer.

Ten patients with small cell lung cancer were treated with high dose human lymphoblastoid interferon (50-100 megaunits m-2) for 5 days, followed by low dose interferon (3 megaunits m-2) for 3 weeks. At the end of treatment, and one month later, there was no evidence of either complete or partial response. The treatment produced fever, anorexia and weight loss, with transient leucopenia and thrombocytopenia; there was evidence of a non-cholestatic elevation of serum alanine aminotransferase, with clinical deterioration in the condition of three patients presenting with hyponatraemia. A transient hypocalcaemia during high dose therapy was also noted. It seems that lymphoblastoid interferon as a single agent is unlikely to have a role in the treatment of small cell lung cancer, and that its administration as employed in this study is associated with considerable toxicity.

Aged↗
PubMed1983

Interactions of DNA replication factors in vivo as detected by introduction of suppressor alleles of dnaA into other temperature-sensitive dna mutants.

Suppressor mutations located within dnaA can suppress the temperature sensitivity of a dnaZ polymerization mutant, indicating in vivo interaction of the products of these genes. The suppressor allele of dnaA [designated dnaA(SUZ, Cs)] could not be introduced, even at the permissive temperature, by transduction into temperature-sensitive (Ts) dnaC or dnaG recipients; it was transduced into dnaB(Ts) and dnaE(Ts) strains but at very low frequency. Recipient cells which were dnaA+ dnaE(Ts) were killed by the incoming dnaA(SUZ, Cs) allele, and it is presumed that combinations of dnaA(SUZ, Cs) with dnaB(Ts), dnaC(Ts), or dnaG(Ts) are lethal also. In one specific case, the lethality required the presence of three alleles: the incoming dnaA suppressor mutation, the resident dnaA+ gene, and the dnaB(Ts) gene. This was shown by the fact that dnaB(Ts) could readily be introduced into a dnaA(SUZ, Cs) dnaB+ recipient. That is, in the absence of dnaA+, the dnaA suppressor and dnaB(Ts) double mutant was stable. One model to explain these results proposes that the dnaA protein functions not only in initiation but also in the replication complex which contains multiple copies of dnaA and other replication factors.

Bacterial Proteins↗
PubMed1983

Physiological properties of cold-sensitive suppressor mutations of a temperature-sensitive dnaZ mutant of Escherichia coli.

Suppressors of a temperature-sensitive dnaZ polymerization mutant of Escherichia coli have been identified by selecting temperature-insensitive revertants. Those suppressed strains which concomitantly became cold sensitive were chosen for further study. Intragenic suppressor mutations, which caused cold-sensitive defects in DNA polymerization, were located in dnaZ by transduction with lambda dnaZ+ phages. Extragenic suppressor mutations were mapped within the initiation gene dnaA. These suppressor-containing strains were defective in initiation at low temperature as determined by measurements of DNA synthesis in vivo and in toluene-treated cells. The occurrence of suppressor mutations of dnaZ(Ts) within the dnaA gene is considered evidence that the dnaA and dnaZ products interact in vivo. A second indication of a dnaA-dnaZ protein-protein interaction was provided by the observation that the introduction of additional copies of the dnaZ+ gene into a strain carrying the dnaA suppressor mutation was lethal [whether the strain was dnaZ+ or dnaZ(Ts)].

Bacterial Proteins↗
PubMed1982

The Escherichia coli dnaW mutation is an allele of the adk gene.

A dnaW mutant, isolated on the basis of inability to effect conjugal DNA transfer at high temperature, has been shown by complementation and enzyme assay to be defective in the adk (adenylate kinase; EC 2.7.4.3) locus. The adk mutant, known to have reduced ATP concentration at the nonpermissive temperature (Cousin and Belaich 1966), was used to demonstrate a donor energy requirement for stable aggregate formation and for chromosome transfer in conjugation.

Adenylate Kinase↗
PubMed1982

Properties of a Herpes virus-transformed hamster cell line--I. Growth and culture characteristics of sublines of high and low metastatic potential.

A Herpes virus hominis type 2-induced tumour line of the Syrian hamster, designated HSV-2-333-2-26, exhibited a low level of spontaneous metastasis from the primary subcutaneous tumour mass. Two lung foci, met A and met B, were resected and transplanted s.c.; both lines showed an elevated metastatic potential compared with the parent cell line such that all animals inoculated with these cells developed metastatic disease within 40 days after resection of the primary tumour. Neither the rapidity of cell growth in vivo or in vitro, nor anchorage-independent growth or degree of attachment to a solid substrate in vitro are implicated in the altered behaviour of the metastatic cell lines.

Animals↗
PubMed1982

Interaction of the Escherichia coli dnaA initiation protein with the dnaZ polymerization protein in vivo.

To define in vivo interactions of Escherichia coli DNA replication components, extragenic suppressors of a dnaZ(TS) mutant were isolated. A temperature-sensitive dnaZ mutant, which is defective in polymerization, was placed at 39 degrees C to select temperature-insensitive revertants. Some of these revertants also were cold sensitive, a phenotypic property that facilitated study of the suppressor. Mapping of the cold sensitivity indicated that some of the suppressor mutations are intragenic but others are located within the initiation gene, dnaA. The dnaA mutations that suppress the dnaZ(TS) defect are designated dnaA(SUZ, CS). The dnaA(SUZ, CS) strains have a defect in DNA synthesis at low temperature that is typical of an initiation defect. These data suggest that the dnaA product, an initiation factor, interacts in vivo with the dnaZ protein, a polymerization factor.

Bacterial Proteins↗
PubMed1982

An immunoradiometric assay of serum interferon using a monoclonal antibody.

An immunoradiometric assay for human interferon-alpha (HuIFN-alpha) has been adapted for the assay of low concentrations of HuIFN-alpha in human serum. The sensitivity of the assay is 5 to 10 IU/ml and the coefficient of variation less than 10%. The assay was shown to compare well with a biological (antiviral) assay in the measurement of serum interferon following intramuscular injection of HuIFN-alpha in nine volunteers. Serum interferon was also measured in the serum from 250 normal human donors. Two donors appeared to have detectable levels of HuIFN-alpha.

Antibodies, Monoclonal↗
PubMed1982

The pharmacology of benoxaprofen with particular to effects on lipoxygenase product formation.

Benoxaprofen has three pharmacological activities which may relate to its clinical activity profile. The most important of these for anti-inflammatory therapy is the regulation of directional monocyte movement in response to a stimulus but the inhibitory activity against lipoxygenase may be of considerable relevance in the treatment of asthma. The relatively weak inhibitory action on prostaglandin synthetase probably confers clinical benefit with a reduced potential to cause severe gastric side effects. Benoxaprofen is an inhibitor pf the lipoxygenase enzyme which converts arachidonic acid to hydroxy derivatives and the leukotrienes. These products are potent pharmacological agents with potentially important roles in inflammation and hypersensitivity disorders.

Anaphylaxis↗
PubMed1981

Metabolic requirements for rabbit polymorphonuclear leucocyte lipoxygenase activity.

Lipoxygenase product formation by rabbit PMNs on exposure to the divalent ionophore A23187 was dependent upon pH, metabolic energy and substrate availability. Formation of 5-HETE and 5,12-diHETE (the major arachidonate oxygenation products) was inhibited by calcium depletion, by the metabolic inhibitors iodoacetate and 2-deoxyglucose, and by the substrate inhibitor TYA. The reaction had a pH optimum of 7.0-7.5. Colchicine, cytochalasin B and isoprenaline had no significant effects on HETE formation.

Animals↗
PubMed1980

The effects of some antirheumatic drugs on an in vitro model of human polymorphonuclear leucocyte chemokinesis.

1 A rapid, reproducible in vitro assay for studying the chemokinetic movement of human polymorphonuclear leucocytes (PMNs) is described. Two synthetic peptides, formyl methionyl-leucyl-phenylalanine (FMLP) and formyl methionyl-phenylalanine (FMP), were used as a standard chemokinesins. 2 Maximal chemokinetic movement was observed with peptide concentrations of 2.5 nM (FMLP) and 100 muM (FMP). EC50 values of 650.0 +/- 60.0 pM and 27.0 +/- 3.5 muM respectively are similar to those reported for chemotactic activity of the peptides in micropore filter assays. 3 The PMN chemokinetic response to FMLP was enhanced by histamine (100 nM) and vitamin C (2.5 muM). 4 Human serum albumin was shown to induce chemokinesis but to antagonize the response to FMLP in a dose-related fashion. Fibrinogen similarly antagonized the cell response to peptide. 5 Levamisole (250 nM to 2.5 muM) significantly potentiated the chemokinetic responses to FMLP and FMP in a dose-related manner. The chemokinetic response to FMLP was unaffected by D-penicillamine (250 muM to 10 mM) while alclofenac (500 muM to 1 mM), salicylic acid (250 muM to 10 mM) and indomethacin (100 muM to 1 mM) caused dose-related inhibition.

Anti-Inflammatory Agents↗