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Biomedical subjects

S Barker

Publications and source records attributed to S Barker.

At least 73 records · Page 4Linked to original sources
PubMed1996

The N-domain of the human CD66a adhesion molecule is a target for Opa proteins of Neisseria meningitidis and Neisseria gonorrhoeae.

Using COS (African green monkey kidney) cells transfected with cDNAs encoding human cell surface molecules, we have identified human cellular receptors for meningococcal virulence-associated Opa proteins, which are expressed by the majority of disease and carrier isolates. These receptors belong to the immunoglobulin superfamily of adhesion molecules and are expressed on epithelial, endothelial and phagocytic cells. Using soluble chimeric receptor molecules, we have demonstrated that meningococcal Opa proteins bind to the N-terminal domain of biliary glycoproteins (classified as BGP or CD66a) that belong to the CEA (CD66) family. Moreover, the Opa proteins of the related pathogen Neisseria gonorrhoeae, responsible for urogenital infections, also interact with this receptor, making CD66a a common target for pathogenic neisseriae. Over 95% of Opa-expressing clinical and mucosal isolates of meningococci and gonococci were shown to bind to the CD66 N-domain, demonstrating the presence of a conserved receptor-binding function in the majority of neisserial Opa proteins.

Animals↗
PubMed1996

Changes in biologic phenotype of human immunodeficiency virus during treatment of patients with didanosine.

Progression to AIDS in patients harboring human immunodeficiency virus type-1 (HIV-1) isolates expressing a syncytium-inducing (SI) phenotype is faster than in those in whom the virus expresses a non-SI (NSI) phenotype. Zidovudine monotherapy does not appear to alter this outcome. To examine the role of didanosine (ddI) monotherapy in phenotype expression, HIV-1 isolates from 73 patients receiving ddI for up to 72 weeks were analyzed. After 12 weeks, the number of isolates expressing an NSI phenotype was 29% higher than at the start of treatment. Patients receiving high-dose ddI (375 mg twice daily) were significantly more likely to express the NSI phenotype at 12 weeks than patients who received low-dose ddI (100 mg twice daily), even after adjustment for phenotype and CD4 cell count at baseline, suggesting that ddI may be selective against the faster-replicating virus. ddI at 375 mg twice daily significantly increases the probability of an NSI phenotype over the short term in patients with advanced HIV disease.

Antiviral Agents↗
PubMed1995

NMR solution structure of the 32-kDa platelet factor 4 ELR-motif N-terminal chimera: a symmetric tetramer.

Native human platelet factor 4 (PF4) is a homotetrameric protein (70 residues/subunit) known for its anticoagulant heparin binding activity. 2D 15N--1H HSQC NMR experiments of native PF4 in solution show the presence of conformational heterogeneity consistent with the formation of asymmetric homo-tetramers as observed in the X-ray crystal structure of both human and bovine PF4. A chimeric mutant of PF4 (called PF4-M2) which substitutes the first 11 N-terminal residues for the first eight residues from homologous interleukin-8 forms symmetric homo-tetramers with essentially the same heparin binding activity as native PF4. The solution structure of PF4-M2 has been investigated by using two- and three-dimensional 1H- and 15N-NMR spectroscopy and NOE-restrained simulated annealing molecular dynamics. As with other members of the CXC chemokine family whose structures are known, the PF4-M2 subunit monomer consists of a mostly hydrophobic, triple-stranded antiparallel beta-sheet onto which is folded an amphipathic C-terminal helix and a less periodic N-terminal domain. Although N-terminal substitution with the less acidic interleukin-8 sequence most affects the quarternary structure relative to native PF4 at the AC and AD dimer interfaces, AB dimer stability is weakened as reflected in reduced equilibrium association binding constants.

Amino Acid Sequence↗
PubMed1995

Effects of the selective protein kinase C inhibitor Ro 31-7549 on human angiotensin II receptor desensitisation and intracellular calcium release.

The mechanism underlying type I angiotensin II (Ang II) receptor (AT1 receptor) desensitisation is unknown. Structural features suggest it may be a substrate for protein kinase C (PKC). The effects of a selective PKC inhibitor, Ro 31-7549, on receptor desensitisation were investigated in CHO cells expressing the human AT1 receptor. Desensitisation was demonstrated with respect to the calcium response to Ang II in Fura-2-loaded cells. Ro 31-7549 had no effect on desensitisation. However, pretreatment with Ro 31-7549 caused a dose-dependent reduction in calcium release from intracellular stores. PKC may therefore act at a locus distal from the receptor itself.

Angiotensin II↗
PubMed1995

Cloning, characterization and expression of a functional mouse ACTH receptor.

A polymerase chain reaction-generated mouse ACTH-R probe was used to screen a mouse genomic library, and a clone of 13kb containing the entire coding sequence was isolated. The coding sequence shows 84% homology with the human gene at the DNA level and encodes a peptide with 89% homology to the human ACTH-R. This gene is expressed as a major transcript of 1.8kb in the mouse adrenal gland. The gene was expressed in HeLa cells and cAMP production in response to either ACTH or alpha-MSH was measured. cAMP increased in an ACTH dose dependent manner suggesting an EC50 of 7 x 10(-10)M ACTH. alpha-MSH was without effect on this receptor. In conclusion we have cloned a mouse ACTH receptor gene and demonstrated for the first time its expression and functional effect in HeLa cells.

Amino Acid Sequence↗
PubMed1995

Quarternary structure amplification of protein folding differences observed in 'native' platelet factor-4.

Platelet factor 4 (PF4), a protein of 70 residues, exists in solution as a distribution of monomer, dimer and tetramer (M-D-T) states in slow exchange on a 600 MHz 1H-NMR chemical shift time scale. Well-resolved Y60 ring proton resonances in each aggregate state allow derivation of M-D-T populations. Under the same set of solution conditions, M-D-T aggregate state distributions vary from prep to prep or with repeated freeze-drying or raising/lowering the solution pH. These variations are not the result of chemical modifications and reflect differences in the strength of subunit associations and therefore folding. Variations are greatest at pH values at or below the pKas of carboxylate groups, supporting the idea that electrostatic interactions modulate PF4 subunit interactions. Treating these distributions as true equilibria results in free energy differences of about 0.5 kcal/mol subunit. Quarternary structure amplifies free energy differences among various folded states of monomer PF4.

Humans↗
PubMed1995

The relationship between the adrenal tissue renin-angiotensin system, internalization of the type I angiotensin II receptor (AT1) and angiotensin II function in the rat adrenal zona glomerulosa cell.

Many data suggest that the elements of the tissue renin-angiotensin system (RAS) in the adrenal cortex are mostly located in the zona glomerulosa. The relationship of this paracrine/autocrine system with the cellular localization of the angiotensin II (AII) receptor has not bee clarified. Using a specific monoclonal antibody (6313/G2) to the first extracellular domain of the type 1 receptor (AT1), we show here that most of the receptor is internalized in the rat glomerulosa cell. This may result from tonic stimulation by the tissue RAS, and consequent permanent receptor occupancy. When viable glomerulosa cells are incubated with 6313/G2, the receptor is transiently concentrated on the cell surface, and aldosterone output is stimulated. This stimulated output is enhanced by neither threshold nor maximal stimulatory concentrations of AII amide, although the antibody does not inhibit AII binding to the receptor. The antibody directly stimulates inositol trisphosphate (IP3) generation, but, while having no intrinsic action on protein kinase C (PKC) activation, significantly inhibits the PKC response to angiotensin II. The data suggest that although the receptor is mostly internalized, recycling to the plasma membrane is constitutive, or regulated by unknown factors. Retention of the AT1 receptor in the membrane is alone enough to allow sufficient G protein interaction to generate maximal steroidogenic effects, through IP3 generation. PKC activation induced by angiotensin II has no bearing on steroidogenesis in the dispersed glomerulosa cell system.

Adrenal Glands↗
PubMed1995

A formula to calculate blood cross-match requirements for early burn surgery in children.

Excessive bleeding from the burn site accompanies burn surgery, and blood transfusions are therefore an essential and expensive ingredient. A formula, based on the total surface area to be involved in surgery (i.e. burn plus donor area) and the patient's total blood volume, has been in use in our hospital for calculating blood cross-match volumes, but has not been scientifically evaluated. Prospectively the predicted blood loss, based on the formula in 111 consecutive paediatric burns undergoing surgery, was compared with the actual blood loss as measured using a gravimetric method. Provided that an accurate surface area to be involved in surgery is interposed into the formula, there was a statistically highly significant correlation between the predicted and actual blood loss. Invoking this simple formula may help in decreasing unnecessary cross-matching, and discarding, of blood products.

Blood Grouping and Crossmatching↗
PubMed1995

Meningococcal pilin: a glycoprotein substituted with digalactosyl 2,4-diacetamido-2,4,6-trideoxyhexose.

Neisseria meningitidis pili are filamentous protein structures that are essential adhesins in capsulate bacteria. Pili of adhesion variants of meningococcal strain C311 contain glycosyl residues on pilin (PilE), their major structural subunit. Despite the presence of three potential N-linked glycosylation sites, none appears to be occupied in these pilins. Instead, a novel O-linked trisaccharide substituent, not previously found as a constituent of glycoproteins, is present within a peptide spanning amino acid residues 45 to 73 of the PilE molecule. This structure contains a terminal 1-4-linked digalactose moiety covalently linked to a 2,4-diacetamido-2,4,6-trideoxyhexose sugar which is directly attached to pilin. Pilins derived from galactose epimerase (galE) mutants lack the digalactosyl moiety, but retain the diacetamidotrideoxyhexose substitution. Both parental (#3) pilins and those derived from a hyper-adherent variant (#16) contained identical sugar substitutions in this region of pilin, and galE mutants of #3 were similar to the parental phenotype in their adherence to host cells. These studies have confirmed our previous observations that meningococcal pili are glycosylated and provided the first structural evidence for the presence of covalently linked carbohydrate on pili. In addition, they have revealed a completely novel protein/saccharide linkage.

Amino Acid Sequence↗