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D A Hopkinson

Publications and source records attributed to D A Hopkinson.

At least 19 recordsLinked to original sources
PubMed1992

Phosphoglucomutase 1: complete human and rabbit mRNA sequences and direct mapping of this highly polymorphic marker on human chromosome 1.

A cDNA clone encoding the mRNA for the highly polymorphic human enzyme phosphoglucomutase 1 (PGM1; EC 5.4.2.2) has been isolated and characterized. This was achieved indirectly by first isolating a rabbit cDNA from an expression library using anti-rabbit PGM antibodies. A comparison of the nucleotide sequences shows that the homologies between human and rabbit PGM1 mRNAs are 92% and 97% for the coding nucleotide sequence and the amino acid sequence, respectively. The derived rabbit amino acid sequence is in complete agreement with the published protein sequence for rabbit muscle PGM. A physical localization of the human PGM1 gene to chromosome 1p31 has been determined by in situ hybridization. Analysis of DNA from a wide variety of vertebrates indicates a high level of PGM1 sequence conservation during evolution.

Animals↗
PubMed1992

Detection of ABO blood group polymorphism by denaturing gradient gel electrophoresis.

We report the use of a polymerase chain reaction (PCR) format together with denaturing gradient gel electrophoresis (DGGE) which allows rapid identification of the 6 major genotypes (AA, AO, BB, BO, AB and OO) of the human ABO blood group polymorphism in a single amplification. The procedure also distinguishes hitherto undescribed polymorphisms associated with the O and B alleles. Thus in testing 95 unrelated European individuals 4 different O alleles, 2 B alleles and 1 A allele were identified by DGGE and the level of recognisable heterozygosity, and hence the information content of the locus as a genetic marker, was raised from 3/95 (3%) to 66/95 (70%). The procedure is robust, genotyping is rapid and clear-cut, and has immediate implications for the use of the ABO locus in linkage analysis on chromosome 9q, the investigation of disease associations and forensic identification.

ABO Blood-Group System↗
PubMed1991

Detection of the common alpha-1-antitrypsin variants by denaturing gradient gel electrophoresis.

The well-characterized and highly polymorphic human alpha-1-antitrypsin (AAT) gene was used as a test locus to evaluate the general applicability of denaturing gradient gel electrophoresis (DGGE) for the detection of single base change polymorphisms. We report the resolution of all the major alleles, M1Ala, M1Val, M2, M3, S and Z and the identification of substantial genetic polymorphism in intron 3-4 by this technique. DGGE was found to be a quick and efficient method for the screening of multiple samples for the presence of genetic variation.

Alleles↗
PubMed1991

Antigenic analysis of the major human phosphoglucomutase isozymes: PGM1, PGM2, PGM3 and PGM4.

The cross-reactivity of human phosphoglucomutase isozymes (PGM1, PGM2, PGM3 and PGM4) has been investigated using anti-rabbit muscle PGM polyclonal antibodies. Significant differences were revealed: an IgG fraction of the antiserum reacted with the primary and secondary PGM1 isozymes of all the common phenotypes. However, there was no reaction with the PGM2 or PGM3 isozymes; thus these latter isozymes share no major antigenic determinants with human or rabbit PGM1 and are therefore structurally distinct. In contrast, the PGM isozymes of human milk attributed to a fourth locus, PGM4, showed similar cross-reactivity as PGM1 suggesting close structural similarity. The IgG was also employed as a reagent to remove PGM1 from haemolysates so as to allow the unambiguous assessment of the PGM2 isozyme patterns by isoelectric focusing. However, no proven genetic variation was encountered in a sample of 32 individuals.

Cross Reactions↗
PubMed1991

Identification of a conserved epitope in class I alcohol dehydrogenase isoenzymes using monoclonal antibodies.

Two monoclonal antibodies raised against native horse alcohol dehydrogenase (HADH) bind preferentially to the enzyme attached to solid supports and recognize the denatured and carboxymethylated HADH subunits. Both antibodies cross-react with the human class I isoenzymes but do not recognize the class III ADH isoenzyme. Protease digestion, electrophoresis and HPLC have been used to identify the linear epitope which is contained in the sequence Pro344-Glu357 of the HADH subunit.

Alcohol Dehydrogenase↗
PubMed1990

HLA phenotypes and gene polymorphisms in juvenile liver disease associated with alpha 1-antitrypsin deficiency.

Chronic liver disease affects up to 20% of children with alpha 1-antitrypsin deficiency owing to the PiZZ genotype. Previous observations of a familial occurrence and abnormal immune responses to liver antigens in these patients suggests that immunoregulatory genes may be involved in the pathogenesis of liver damage. We have identified HLA phenotypes and class II (HLA-DR) gene polymorphisms in 140 white PiZZ subjects, of whom 92 (83 index patients) had liver disease, and 206 first-degree relatives. DR3* was present in 35 of 75 (46.7%) unrelated patients with liver disease compared with 5 of 28 (17.8%) patients without (p less than 0.01) and 23 of 100 controls (p less than 0.001). DR4 was increased in patients without liver disease; it was present in 17 of 28 (60.7%) compared with 29 of 75 (38.7%) patients with liver disease (p less than 0.05) and 36 of 100 controls (p less than 0.025). Using Southern blot analysis with HLA-DRB and DQB DNA probes, we identified two polymorphisms of DR3, only one (Dw25) of which is raised in PiZZ individuals with liver disease (9 of 55: 16.4%) compared with 1 of 23 (4.4%) without and 2 of 52 (3.9%) controls (p less than 0.05). Analysis of the segregation of HLA haplotypes in 77 families revealed no concordance for liver disease with HLA in those with affected sibships, indicating that, although DR3-Dw25 is associated with liver disease in alpha 1-antitrypsin deficiency, other factors must play a pathogenic role.

Adolescent↗
PubMed1990

Pyrimidine 5'-nucleotidase activity in normal and deficient human lymphoblastoid cells.

Two distinct pyrimidine 5'-nucleotidases (UMPH-1 and UMPH-2) have previously been detected in human erythrocytes; UMPH-1 is deficient in a haemolytic anaemia, while UMPH-2 is unaffected. Only the erythrocyte shows pathological effects in this disorder. Here we have studied lymphoblastoid cell lines from control and UMPH-1 deficient patients to determine whether UMPH-1 can be detected in lymphoblastoid cells and whether the deficiency of UMPH-1 results in any measurable metabolic effects. Both UMPH-1 and a UMPH-2-like activity were found to be present in control lymphoblastoid cells. UMPH-1 was undetectable in the patients' cells; minor but significant changes were found in the pyrimidine nucleotide and nucleoside pools in the cells.

5'-Nucleotidase↗
PubMed1990

Inosine triphosphate pyrophosphohydrolase deficiency in a kindred with adenosine deaminase deficiency.

A complete deficiency of inosine triphosphate pyrophosphohydrolase (ITPase) has been identified, together with high concentrations (mean 157 mumol/l) of the unusual nucleotide ITP, in the erythrocytes of 3 members of a consanguineous United Kingdom kindred. The defect has been noted previously in North America and Sweden, but even in presumed homozygotes some residual ITPase activity was reported. Homozygosity for the defect has not been associated previously with any clinical abnormality. In this kindred it was co-existent with adenosine deaminase (ADA) deficient severe combined immunodeficiency. Since the genes for both ITPase and ADA are localised on the same chromosome, segregation analysis of ITPase and ADA activity was undertaken in available kindred members. The results confirmed an autosomal recessive mode of inheritance for ITPase deficiency, but suggested that the co-existence with ADA deficiency was coincidental.

Adenosine Deaminase↗
PubMed1989

Variation in alpha-1-antitrypsin phenotypes associated with penicillamine therapy.

During a detailed study of alpha-1-antitrypsin (AAT) by isoelectric focusing, of 130 individuals of the PiZ phenotype and their families, an unusual alpha-1-antitrypsin protein pattern was identified which related directly to the effect of penicillamine therapy. The effect, believed to be mediated by reaction with the free cysteine of the alpha-1-antitrypsin molecule, was demonstrable by in vitro experiments. It was not apparently associated with loss of the anti-proteinase properties of alpha-1-antitrypsin and was not specific to the Z gene products. It is however, a source of possible confusion in the accurate assessment of Pi phenotypes.

Cystamine↗
PubMed1989

Isozyme and DNA analysis of human S-adenosyl-L-homocysteine hydrolase (AHCY).

Erythrocyte and tissue isozymes of human AHCY have been studied by starch gel electrophoresis, cellulose acetate electrophoresis, isoelectric focusing and Na dodecyl sulphate electrophoresis. The same isozyme was observed in all the tissues studied, suggesting that human AHCY is encoded by a single structural locus. Two variant alleles were identified in erythrocyte AHCY using starch gel electrophoresis in a sample of 166 unrelated individuals from the British population. The gene frequencies were 0.024 for AHCY*2 and 0.006 for AHCY*3. The variant isozyme patterns could not be distinguished by isoelectric focusing. Using the homologous rat cDNA AHCY probe, human AHCY cDNA recombinants were isolated from a placental cDNA library. The human and rat sequences show considerable homology in the coding region of the gene and also, but to a lesser extent, in the distal part of the 3' untranslated region. Preliminary observations suggest the occurrence of a high frequency PvuII site RFLP identified with the human AHCY probe.

Adenosylhomocysteinase↗
PubMed1989

Genetic studies on a new deficiency gene (PI*Ztun) at the PI locus.

During a study of the alpha 1 antitrypsin (AAT) protein and its locus (PI) by high resolution isoelectric focusing and direct molecular analysis of 106 PIZ probands and their families, a new allele (Ztun) was identified that resembles Z in many of its properties. Two sibs, both compound heterozygotes for Ztun and Z, showed similar evidence of mild liver involvement that was indistinguishable from that associated with classical ZZ homozygotes. The Ztun protein appeared to be deficient in the plasma to about the same degree as the Z protein. Allele specific oligonucleotide analysis of amplified genomic DNA indicated that the new allele is the result of a mutation in exon V that is identical to the classical G----A transition at codon 342 that results in the Glu----Lys substitution characteristic of the Z allele. An analysis of DNA haplotypes constructed from polymorphic restriction enzyme recognition sites in and around the PI locus confirmed that Ztun probably represents a new mutation at codon 342 that has occurred on an M2-like genetic background.

Alleles↗
PubMed1989

Rhodanese isozymes in three subjects with Leber's optic neuropathy.

Previous studies have reported an association between Leber's optic neuropathy and deficiency of rhodanese activity in liver and rectal mucosa. We have studied the rhodanese isozymes in liver biopsies from three subjects with Leber's optic neuropathy. The rhodanese isozyme patterns were indistinguishable from controls both in relative intensity and position on the isoelectric focusing pH gradient. No new rhodanese isozymes were observed and there was no evidence of deficiency in any of the cases.

Adult↗
PubMed1987

Studies of genetic linkage between adult polycystic kidney disease and three markers on chromosome 16.

Adult polycystic kidney disease (APKD) is a common genetic disorder that is inherited as an autosomal dominant trait. Recent reports show that, in some families, the APKD gene shows close genetic linkage to two chromosome 16 specific genetic markers. We have been conducting a genetic linkage study using 29 polymorphic isoenzyme and antigenic markers in 184 members of 12 APKD families. We present here the results of linkage analysis using three of these markers which have also been reported to be located on chromosome 16: phosphoglycolate phosphatase (PGP), glutamate pyruvate transaminase (GPT), and haptoglobin (HP). The results show that APKD is closely linked to the PGP locus on the short arm of chromosome 16 (16p13----p12), which is consistent with the previously reported linkage both to PGP and to the alpha globin locus. The genetic distance between PGP and APKD shows a maximum likelihood value of the recombination fraction at zero with a lod score of 5 X 5. There is no evidence of linkage between APKD and either GPT or HP. The PGP polymorphism potentially provides a useful predictive test to complement the use of alpha globin probes in genetic counselling. These tests should provide an efficient means of primary screening of family members at risk, as well as introducing the possibility of prenatal diagnosis.

Alanine Transaminase↗
PubMed1986

Two genetic markers closely linked to adult polycystic kidney disease on chromosome 16.

The genetic locus for autosomal dominant adult polycystic kidney disease was recently assigned to chromosome 16 by the finding of genetic linkage to the alpha globin gene cluster. Further study showed that the phosphoglycolate phosphatase locus is also closely linked to both the locus for adult polycystic kidney disease and the alpha globin gene cluster. These findings have important implications for the prenatal and presymptomatic diagnosis of adult polycystic kidney disease and for a better understanding of its pathogenesis.

Adolescent↗