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Biomedical subjects

T Wilkinson

Publications and source records attributed to T Wilkinson.

At least 19 recordsLinked to original sources
PubMed1995

Complex formation between the NS3 serine-type proteinase of the hepatitis C virus and NS4A and its importance for polyprotein maturation.

Processing of the hepatitis C virus polyprotein is mediated by host cell signalases and at least two virally encoded proteinases. Of these, the serine-type proteinase encompassing the amino-terminal one-third of NS3 is responsible for cleavage at the four sites carboxy terminal of NS3. The activity of this proteinase is modulated by NS4A, a 54-amino-acid polyprotein cleavage product essential for processing at the NS3/4A, NS4A/4B, and NS4B/5A sites and enhancing cleavage efficiency between NS5A and NS5B. Using the vaccinia virus-T7 hybrid system to express hepatitis C virus polypeptides in BHK-21 cells, we studied the role of NS4A in proteinase activation. We found that the NS3 proteinase and NS4A form a stable complex when expressed as a single polyprotein or as separate molecules. Results from deletion mapping show that the minimal NS4A domain required for proteinase activation is located in the center of NS4A between amino acids 1675 and 1686 of the polyprotein. Amino acid substitutions within this domain destabilizing the NS3-NS4A complex also impair trans cleavage at the NS4A-dependent sites. Similarly, deletion of amino-terminal NS3 sequences impairs complex formation as well as cleavage at the NS4B/5A site but not at the NS4A-independent NS5A/5B site. These results suggest that a stable NS3-NS4A interaction is important for cleavage at the NS4A-dependent sites and that amino-terminal NS3 sequences and the central NS4A domain are directly involved in complex formation.

Algorithms↗
PubMed1991

Hb Geelong [beta 139(H17)Asn----Asp].

Hb Geelong [beta 139(H17)Asn----Asp] was detected in a German woman of Polish-Russian descent. It is an unstable variant which appears to increase the severity of a beta (+)-thalassemic phenotype in the propositus. The electrophoretic properties of Hb A and Hb Geelong are similar on cellulose acetate in both acidic and alkaline conditioning. The electrophoretic mobility and the amino acid analysis of beta XT-14 indicated the substitution Asn----Asp at beta 139. The sequence of beta XT-14 was confirmed by dansyl-Edman degradation. The slight increase observed in the P50 of whole blood is not intrinsic to the beta 139 substitution, but is thought to result from an increased 2,3-diphosphoglycerate level in response to anemia. No family studies were possible to investigate the mode of inheritance of either beta (+)-thalassemia or Hb Geelong in the propositus. Synthetic globin chain ratios suggest that impaired synthesis of the variant globin chain is partially responsible for the low level of Hb Geelong in peripheral blood.

2,3-Diphosphoglycerate↗
PubMed1990

Molecular and hematologic characterization of Scottish-Irish type (epsilon gamma delta beta)zero thalassemia.

The DNA deletion associated with an example of (epsilon gamma delta beta)zero thalassemia (Scottish-Irish type) was characterized. The deletion is approximately 205 kb in length and involves the epsilon, G gamma, A gamma, delta, and beta globin genes. The breakpoint is located 263 bp 3' to exon 3 of the beta globin gene. An LI (KpnI) repeat element approximately 320 bp in size is found at the 3' end of the novel DNA sequence. Different clinical phenotypes for three heterozygous neonates suggest that the deletion alone does not predict severity of (epsilon gamma delta beta)zero thalassemia at this age.

Adolescent↗
PubMed1989

The anatomic relationship of the insertion of the superior lateral pterygoid muscle to the articular disc in the temporomandibular joint of human cadavers.

Confusion has existed as to whether the major insertion of the superior lateral pterygoid muscle is into the articular disc or the condyle. Five human cadaver joints were studied under the dissecting microscope. This allowed superior lateral pterygoid fibres to be selectively placed under tension to determine their point of insertion and to examine the integrity of the anterior joint capsule. All superior lateral pterygoid fibres gained either direct or indirect insertion to the condyle. A classification for the variation in insertion is suggested. The fibres of the anterior joint capsule extended from the anterior rim of the condyle to the roof of the infratemporal fossa but under the foot of the disc they blended with and became indistinguishable from disc fibres.

Cadaver↗
PubMed1986

Molecular defects in 2 examples of severe Hb H disease.

Severe Hb H disease presented in unexpected ways in 2 families of Greek origin. In 1, Hb H disease led to neonatal death. The underlying molecular defect was double-heterozygosity for the --Med/ alpha thalassaemia haplotype and a nondeletional alpha thalassaemia defect (alpha alpha T'Karditsa'/). The 2nd family requested antenatal diagnosis. The husband had mild nondeletional alpha thalassaemia. Initial investigations in the wife demonstrated unexpected gene mapping patterns. These have recently been shown to result from the (-alpha)Med 20.5/ haplotype.

Chromosome Mapping↗
PubMed1986

Alpha thalassemia British type (alpha alpha/--Brit) in an Australian family.

Alpha thalassemia is rarely diagnosed in Australian families of British or Northern European ancestry. In 1972, a third generation Australian was shown to have alpha thalassemia. In the absence of known Mediterranean or South East Asian ancestry it was reported as being the first example of alpha thalassemia in an Australian family. Further study of the proposita in 1985 using DNA mapping of the alpha globin gene complex, shows a distinctive molecular defect identical to the British type of alpha thalassemia. The latter is clearly different from the commonly encountered Mediterranean and South East Asian alpha zero haplotypes. Recognition that alpha zero thalassemia occurs in Australians is important since it may produce a microcytic hypochromic anemia. Its inheritance together with other forms of alpha thalassemia may lead to severe Hb H disease or Hb Bart's hydrops fetalis.

Adult↗
PubMed1986

Globin genes in Polynesians have many rearrangements including a recently described gamma gamma gamma gamma/.

Rearrangements involving genes of the alpha- and beta-globin loci were frequently detected in DNA from Polynesians. A founder effect and genetic drift occurring 2,000-3,000 years ago as Polynesians migrated eastward across the Pacific is proposed as the likely mechanism for these genetic changes that include deletions or additions of alpha-, gamma-, and zeta-globin genes and an unusual restriction fragment length polymorphism (RFLP) associated with the zeta gene. Preliminary data show different frequencies for gene rearrangements between island groups. Further study of these differences should provide additional information on the prehistory of Polynesians.

Chromosome Mapping↗
PubMed1984

Identification of beta variant hemoglobins by DNA restriction endonuclease mapping.

An alternative method for identifying beta variant hemoglobins is described. Computer analysis of restriction sites was used to predict which beta variants could be detected by DNA mapping. 61 of 217 variants were shown to have changes in restriction fragment patterns which were useful markers for the abnormal hemoglobin. A further 25 could be identified by polyacrylamide electrophoresis. Implications of DNA analysis in diagnosis of variant hemoglobins are discussed.

Base Sequence↗
PubMed1983

Equilibria amongst different molybdenum (V)-containing species from sulphite oxidase. Evidence for a halide ligand of molybdenum in the low-pH species.

The interaction of chloride, fluoride and phosphate ions with the molybdenum centre of sulphite oxidase in the pH range 6.2 to 9.6 has been studied by e.p.r. of Mo(V) in the enzyme reduced by sulphite. Detailed studies were made from e.p.r. spectra recorded at about 120K and more limited studies from spectra of liquid samples at about 295K and also from enzyme activity measurements. Interconversion between low-pH and high-pH Mo(V) e.p.r. signal-giving species [described by Lamy, Gutteridge & Bray (1980) Biochem. J. 185, 397-403] is influenced by chloride concentration, a 10-fold increase in concentration (in the range of about 1 mM to 100 mM) causing an increase of about 1 pH unit in the apparent pK for the conversion. This suggests that chloride is a constituent of the low-pH species. In support of this, high concentrations of fluoride modified the e.p.r. spectrum. Partial conversion to a Mo(V) species, in which F- has presumably replaced Cl- and showing hyperfine coupling of A(19F)av. 0.5mT, is indicated. It is proposed that interconversion between high-pH and low-pH species is of the form: (formula; see text) No evidence that Cl- is essential for enzymic activity was found. Data relating to equilibria amongst low-pH, high-pH and also the phosphate species are presented. Depending on pH and on concentrations of Cl- and H2PO4-, one, two, or all three species may be present. Qualitatively, under appropriate conditions, the phosphate species tends to replace some or all of the low-pH species. Quantitative analysis by a computer procedure permitted an appropriate scheme to be deduced and equilibrium constants to be evaluated. Studies on the e.p.r. signals at 295K indicated that similar equilibria applied in liquid solution, but with some changes in the values of the constants. The structure of the molybdenum centre in its various states and the nature of the enzymic reaction are discussed.

Chlorides↗
PubMed1983

A new hemoglobin with high oxygen affinity--hemoglobin bunbury: alpha 2 beta 2 [94 (FG1) Asp replaced by Asn].

Hemoglobin Bunbury was detected in an Italian woman from Bunbury, Western Australia, following detection of an abnormal hemoglobin by electrophoresis. The variant form accounted for 38% of the total hemoglobin and migrated on cellulose acetate electrophoresis between HbS and HbA. Structural analysis demonstrated a new substitution in the beta chain, beta 94 Asp replaced by Asn. Hemoglobin Bunbury is stable to both heat and isopropanol denaturation. This variant shows an increase in oxygen affinity and a reduced Bohr effect. Heterozygosity for Hb Bunbury produces no clinical symptoms or compensatory erythrocytosis.

Adult↗
PubMed1982

Evidence from electron-paramagnetic-resonance spectroscopy for a complex of sulphite ions with the molybdenum centre of sulphite oxidase.

Reduction of sulphite oxidase by sulphite at low pH values in Mes (4-morpholine-ethanesulphonic acid) buffer gives rise to a new molybdenum(V) electron-paramagnetic-resonance spectrum different from that obtained by photoreduction of the enzyme in the same medium. The spectrum is attributed to a sulphite complex of the enzyme, showing g-values of about 2.000, 1.972 and 1.963. The complex is analogous to that with the inhibitor phosphate in that it gives rise to no observable hyperfine coupling of Mo(V) to exchangeable protons.

Catalysis↗