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Biomedical subjects

J Pumphrey

Publications and source records attributed to J Pumphrey.

12 recordsLinked to original sources
PubMed2000

Serological prevalence of herpes simplex virus type 2 amongst GUM clinic attenders in a district general hospital setting.

Our objective was to determine the seroprevalence of herpes simplex virus (HSV) type 2 infection amongst genitourinary medicine (GUM) clinic attenders at a district general hospital using a commercially available enzyme immunoassay (EIA). In a prospective study, heterosexual patients attending the Department of GUM at Trafford General Hospital attending with a new clinical problem and having a blood sample taken for routine syphilis serology had the same sample tested for HSV type 2 antibodies. The prevalence of HSV type 2 seropositivity amongst participants was 9.9% (24/242) for men and 18.7% (46/246) for women. With respect to undiagnosed, asymptomatic infection the seroprevalence was 8.6% and 17% respectively. For those attenders locally resident the seroprevalence was 10.1% and 17.5% respectively, and undiagnosed, asymptomatic infection 8.5% and 17.1% respectively. Although seroprevalence figures in this study are lower than the only previous report in the UK, these results, nevertheless, show that seropositivity is not confined to large urban centres. Patients attending GUM clinics are likely to have high rates of undiagnosed HSV type 2 infection.

Antibodies, Viral↗
PubMed1997

Application of 'Clearview Chlamydia' for the rapid detection of cervical chlamydial antigen.

In a study to evaluate the Clearview chlamydia test in a genitourinary medicine clinic, the sensitivity and specificity were 76.4% and 99.4% respectively when compared with the DAKO IDEIA chlamydia test processed in a department of medical microbiology. The time taken to perform and read the Clearview chlamydia test 'on site' did not interfere with the normal routine functioning of the clinic and the nurses who performed it as part of their routine duties found it simple and easy to use.

Chlamydia Infections↗
PubMed1990

Sequence and structure of Clp P, the proteolytic component of the ATP-dependent Clp protease of Escherichia coli.

The ATP-dependent Clp protease of Escherichia coli contains two dissimilar components: the Clp A regulatory polypeptide, with two ATP binding sites and intrinsic ATPase activity, and the Clp P subunit, which contains the proteolytic active site. The DNA sequence of the clpP gene predicts a protein of 207 amino acids (Mr 21,679), which is in close agreement with the size determined by sodium dodecyl sulfate-gel electrophoresis of purified Clp P. Clp P has a native Mr of approximately 240,000, and electron micrographs of the protein show superimposed disk-like structures with a central cavity, similar in appearance to purified proteasomes from eukaryotic cells. Clp P is synthesized with a 14-amino acid leader which is rapidly cleaved in vivo to yield the 193-amino acid protein which has activity in vitro. The clpP gene is at 10 min on the E. coli map, close to that for the ATP-dependent Lon protease of E. coli and far from the gene for clpA. Primer extension experiments indicate that transcription initiates immediately upstream of the coding region for Clp P, with a major transcription start at 120 bases in front of the start of translation. Insertion mutations in clpP have been isolated and transferred to the chromosome; strains devoid of Clp P are viable in the presence or absence of Lon protease. Mutations in clpP stabilize the same Clp A-beta-galactosidase fusion protein specifically stabilized by clpA mutations, providing the first genetic evidence that Clp A and Clp P act together in vivo.

ATP-Dependent Proteases↗
PubMed1988

The two-component, ATP-dependent Clp protease of Escherichia coli. Purification, cloning, and mutational analysis of the ATP-binding component.

The ATP-binding component (Component II, hereafter referred to as ClpA) of a two-component, ATP-dependent protease from Escherichia coli has been purified to homogeneity. ClpA is a protein with subunit Mr 81,000. It has an intrinsic ATPase activity and activates degradation of protein substrates only in the presence of a second component (Component I, hereafter referred to as ClpP), Mg2+, and ATP. The amount of ClpA varies by less than a factor of 2 in cells grown in different media and at temperatures from 30 to 42 degrees C. ClpA does not appear to be a heat-shock protein since its synthesis is not dependent on htpR. Antibodies against purified ClpA were used to identify lambda transducing phage bearing the clpA gene. The cloned gene contains a DNA sequence expected to code for the first 28 amino acids of ClpA, which were determined by protein sequencing of purified ClpA. The clpA gene in the phage was mutated by insertion of delta kan defective transposons and the mutations were transferred to E. coli by homologous recombination. The clpA gene was mapped to 19 min on the E. coli chromosome. Mutant cells with insertions early in the gene produce no ClpA protein detectable in Western blots, and extracts of such mutant cells have no detectable ClpA activity. clpA- mutants grow well under all conditions tested and are not defective in turnover of proteins during nitrogen starvation nor in the turnover of such highly unstable proteins as the lambda proteins O, N, and cII, or the E. coli proteins SulA, RcsA, and glutamate dehydrogenase. The degradation of abnormal canavanine-containing proteins is defective in clpA mutants especially in cells that also have a lon- mutation. Extracts of clpA- lon- cells have ATP-dependent casein degrading activity.

ATP-Dependent Proteases↗
PubMed1985

Neoplastic transformation of mast cells by Abelson-MuLV: abrogation of IL-3 dependence by a nonautocrine mechanism.

Normal mast cells can be propagated in culture when medium is supplemented with interleukin-3 (IL-3). We demonstrate that Abelson-MuLV (Ab-MuLV) infection of mast cells eliminates dependence on IL-3 for growth. By contrast, Harvey, BALB, and Moloney MSV, which also productively infect mast cells, are unable to relieve IL-3 dependence. Ab-MuLV-induced IL-3-independent lines express the v-abl-specific transforming protein and have phenotypic characteristics of mast cells. These cells also possess high cloning efficiencies in soft agarose and are tumorigenic in nude mice. In addition, Ab-MuLV induces transplantable mastocytomas in pristane-primed adult mice resistant to lymphoid transformation, defining a new hematopoietic target for malignant transformation by this virus. None of the Ab-MuLV-derived transformants express or secrete detectable levels of IL-3 nor is their growth inhibited by anti-IL-3 serum. These results argue that Ab-MuLV abrogation of the IL-3 requirement is not due to an autocrine mechanism.

Abelson murine leukemia virus↗
PubMed1985

Isolation and expression of complementary DNAs encoding the human interleukin 2 receptor.

Complementary DNAs corresponding to the human receptor for interleukin 2 (IL-2) have been molecularly cloned, sequenced, and expressed in COS-1 cells. The human genome appears to contain a single structural gene for this receptor; however, when transcribed at least two messenger RNAs (mRNAs) are produced which vary in length due to the use of different polyadenylation signals. Sequence analysis of the cloned complementary DNAs indicates an alternate pathway of mRNA processing for this receptor. Splicing of a 216 base pairs segment contained within the protein coding region results in an mRNA unable to code for the IL-2 receptor. In contact complementary DNAs corresponding to unspliced mRNA encode membrane receptors which bind both IL-2 and anti-Tac (monoclonal anti-IL-2 receptor antibody). Analysis of the deduced amino acid sequence reveals that the receptor is composed of 272 amino acids including a signal peptide 21 amino acids in length. Hydrophobicity analysis suggests a single 19 amino acid transmembrane domain. A short intracytoplasmic domain composed of 13 amino acids is present at the carboxy terminus and contains three potential phosphate acceptor sites (serine and threonine but not tyrosine) and typical positively charged amino acids presumably involved in cytoplasmic anchoring. Two sites for N-linked glycosylation sites and numerous extracytoplasmic O-linked glycosylation sites are present.

Amino Acid Sequence↗
PubMed1984

Somatic diversification of immunoglobulins.

A series of three IgM, kappa monoclonal antibodies arising from a fusion of BALB/c spleen cells from mice immunized with beta-(1,6)-galactan-containing antigens have been analyzed. These three lines were found (i) to have homologous protein sequences in the heavy chain D region and at the sites of recombination between the heavy chain variable and D segment (VH-D) and the D and joining segment (D-JH), although amino acid substitutions were observed in both the heavy and light chain variable regions; (ii) to use identical heavy and light chain joining segments; and (iii) to demonstrate two identical (productive and nonproductive) kappa-chain rearrangements. A likely explanation for these observations is that the three lines are clonally related (arise from a common precursor) and that the observed heavy and light chain variable segment substitutions represent somatic point mutations. Because these antibodies are all of the IgM class, the results indicate that a somatic mutational mechanism is activated early in B-cell ontogeny and operates at both the heavy and light chain loci. Furthermore, the somatic mutation process appears to continue during the development of a given cell line, but is independent of class switching.

Amino Acid Sequence↗
PubMed1984

Monoclonal antibodies to DNA and RNA from NZB/NZW F1 mice: antigenic specificities and NH2 terminal amino acid sequences.

Two anti-DNA hybridoma autoantibodies ( A52 , D42 ) were prepared by fusing spleen cells from unimmunized NZB/NZW F1 female mice with BALB/c myeloma cells. The monoclonal antibodies were purified to homogeneity and were analyzed for their antigen-binding specificities. The two anti-DNA antibodies bound single-stranded, double-stranded, and supercoiled DNA, with a marked preference for the single-stranded conformation. Competition experiments performed with synthetic polynucleotides, as well as chain reconstitution experiments, indicated that both the sugar-phosphate backbone and the heterocyclic bases of the nucleic acid are essential for antibody recognition. Amino terminal sequence analysis of A52 and two RNA-binding hybridoma proteins revealed that the heavy chains from all three were members of the VHII subgroup and that the A52 light chain was homologous to the VK8 subgroup. The D42 heavy chain was found to be similar to a phosphocholine-binding hybridoma of the VHIII subgroup.

Amino Acid Sequence↗
PubMed1983

Galactan-binding antibodies. Diversity and structure of idiotypes.

A group of eight IgM hybridoma proteins induced with beta(1,6)-D-galactan-containing antigens has been characterized in terms of primary amino acid sequence and idiotype expression. The H chain amino acid sequences reveal very strong homology in the VH segment although several substitutions are seen that suggest the occurrence of somatic mutation in these IgM molecules. Significant sequence variation was observed in CDR-3, the region generated by the D segment, and the two recombination events, VH-D and D-JH. The number of amino acids in this region contributed by the D segment was found to vary from two to six, yet the overall length of CDR-3 was precisely maintained by the addition of amino acids on either side of D during the recombination processes. These additional amino acids are suggested to result from nucleotide addition by repair enzymes. Idiotypic analysis of these proteins, in conjunction with an assessment of the H chain sequences, has permitted an identification of the molecular basis of both cross-reacting and unique idiotypic determinants expressed by these molecules.

Amino Acid Sequence↗
PubMedDate not supplied

Characterization of idiotypes on antibodies to galactan.

The correlation between the primary structure and the expression of idiotypy in the anti-galactan antibody system is extremely precise. Amino-acid residues, which are implicated in the expression of recurrent or private idiotopes, are localized in the CDR3 region of all hybridomas and of at least 2 out of 4 myeloma proteins. A single substitution in this region may, in some cases, suffice to abolish idiotope expression.

Amino Acid Sequence↗
PubMedDate not supplied

Molecular cloning and expression of cDNAs for the human interleukin-2 receptor.

We have purified the human T-cell growth factor (interleukin-2) receptor and have cloned, sequenced and expressed cDNAs corresponding to this receptor. We identify one gene, but two interleukin-2 receptor mRNAs which differ in their polyadenylation signals. We have isolated an additional cDNA that may correspond to an alternatively spliced mRNA that lacks a 216 base segment and appears to encode an altered membrane protein which cannot bind interleukin-2.

Amino Acid Sequence↗