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Biomedical subjects

R Barker

Publications and source records attributed to R Barker.

At least 73 records · Page 4Linked to original sources
PubMed1986

The structure and transcription start site of a major potato tuber protein gene.

We have isolated recombinant lambda clones containing intact major tuber protein (patatin) genes and flanking sequences from the commercial tetraploid variety Maris Piper. The gene is composed of seven exons and six introns, spread over 4 kb of DNA. Nuclease mapping defined the 5' end of the mRNA approximately 45 bp upstream of the initiation codon. The 5' end of the gene is preceeded by a canonical TATA box sequence. The three known patatin genes encode proteins of nearly identical Mr but very different isoelectric points. The sequence of the gene does not indicate a role for patatin as one of the globulin class of plant storage proteins.

Amino Acid Sequence↗
PubMed1986

Substance P and Parkinson's disease: a causal relationship?

Parkinson's disease (PD) is a common condition that is thought to result from a marked degeneration of dopaminergic neurones of midbrain origin. Here I present evidence to show that PD may result from a primary loss of active tachykinin, probably substance P (SP) in the substantia nigra (SN), and that this loss leads to a secondary degeneration of the dopaminergic neurones. This raises the possibility of treating and curing patients with PD by giving them SP agonsts.

Animals↗
PubMed1986

Effect of inositol hexaphosphate on the transient behavior of red cells following a DMSO-induced osmotic pulse.

An osmotic pulse can be used to incorporate inositol hexaphosphate (IHP) into red cells. The pulse is induced by equilibrating a red cell suspension with DMSO and then rapidly diluting with an isotonic IHP solution. Since IHP binds to hemoglobin and lowers the affinity for oxygen, this method may find application in the preparation of low-affinity cells for experimental and clinical use. The experiments reported here examined the dynamic changes of several red cell variables immediately following the osmotic pulse. The effect of IHP, which has been shown to dissociate red cell cytoskeletons, was evaluated by comparison with a matched phosphate-buffered saline (PBS) diluent. Red cell morphology, volume, and hemoglobin permeability were studied by fixing the cells at times ranging from 0.06 to 300 sec after dilution. Mechanical fragility was measured by subjecting the cells to a short period of shear stress at the same times after dilution. With both diluents, the cells underwent a rapid increase in volume followed by a return towards normal volume with a maximum at less than 250 msec. With IHP diluent, the period of hemoglobin permeability immediately followed the size peak and was completed by about 1 sec after dilution. PBS also induced a second leakage at longer times (10-120 sec), which resulted in a morphological dichotomy with ghosts and intact cells. The choice of diluent also affected sensitivity to shear stress. The IHP-treated cells had a mechanical fragility maximum at about 1 sec. The PBS-treated cells exhibited no enhanced mechanical fragility. An unexpected result was the inhibition of the second phase of lysis in PBS-treated cells by a properly timed shear stress.

Cell Membrane Permeability↗
PubMed1986

Measles virus haemagglutinin gene: cloning, complete nucleotide sequence analysis and expression in COS cells.

A measles virus (Hallé strain) cDNA library was prepared by cloning virus-induced mRNA directly into the expression vector PCD. Clones corresponding to the measles virus haemagglutinin (HA) gene were isolated and one, PCD-HA-15, which corresponded to the complete mRNA sequence, was further characterized. After transfection into COS-7 cells, measles virus HA antigen was detected by immunofluorescence. The [35S]methionine-labelled HA protein from transfected cells was immunoprecipitated by both polyclonal and monoclonal measles virus antibodies. Analysis by SDS-polyacrylamide gel electrophoresis revealed that the PCD-HA-15 protein migrated in a manner identical to the virus-induced HA. Nucleotide sequence analysis established that the gene contained 1949 nucleotides [exclusive of poly(A)] and coded for a protein containing 617 amino acids. A single hydrophobic domain likely to represent the transmembrane region was identified at the N-terminus. A second overlapping reading frame coded for a protein containing 70 amino acids. This contained a short hydrophobic region (16 amino acids) and had two potential N-glycosylation sites. Comparison of the HA gene of the Hallé strain with the published sequence of the Edmonston strain showed that there was a high degree of conservation (99.3%).

Amino Acid Sequence↗
PubMed1984

The effects of physician communications skills on patient satisfaction; recall, and adherence.

An understanding of means to improve patient adherence to the therapeutic regimen is a subject of increasing concern in medical care. This study examined the effects of physician interpersonal skills and teaching on patient satisfaction, recall, and adherence to the regimen. We studied the ambulatory visits of 63 patients to five medical residents at a teaching hospital in Baltimore. It was found that quality of interpersonal skills influenced patient outcomes more than quantity of teaching and instruction. Secondary analyses found that all the effects of physician communication skills on patient adherence are mediated by patient satisfaction and recall. These findings indicate that the physician might pay particular attention to these two variables in trying to improve patient adherence, and that enhancing patient satisfaction may be pivotal to the care of patients with chronic illness.

Adult↗
PubMed1984

System for DNA sequencing with resolution of up to 600 base pairs.

A system capable of resolving about 500 bases is of interest for sequencing of longer DNA molecules. Studies on further optimization of resolution on DNA sequencing gels were carried out. The effect of physico-chemical properties of gels and buffers on resolution were tested, e.g. ionic strength and pH of buffers, different buffer systems, acrylamide concentration, crosslinker concentration, type of crosslinker, temperature of polymerization, denaturing conditions, gel length and thickness. Tested were as well different running conditions like electric field, gel temperature, dimension of sample slots. Gels 0.1-0.2 mm thick and up to 1.2 m long were cast and tested routinely. Gel lengths of 60-70 cm (for sequencing up to 350-400 bases) to about 100 cm (above 400 bases) are practicable. Little is gained in resolution by increasing the gel length from 1 to 1.2 m. Resolution was improved using 0.1 mm thick gels, at a higher pH value of 8.6-8.8, and molarity increased to 0.2 M. The sequencing pattern in the region of higher bases could be better resolved on a twice-magnified picture of that region on the autoradiogram. With the long gels (70-120 cm), it is advantageous to obtain the sequence overlap by running in parallel gels of different concentrations, without re-application of samples, all loaded at the same time. Buffer chamber for running of two of three gels and thermostating plates up to 1.2 m long were designed. In this way four to six thermostated gels can be run from a power supply with two inputs. Three 1 m long gels (concentrations: 4%, 6%, 12-16%) are loaded with several samples of DNA to be sequenced and run in parallel without re-application of the samples. With good samples, the sequence overlap from the gels could be counted up to 500 base pairs, with exceptionally good samples closer to 600 bases. At present this number seems to be near the limit of the resolving power of the polyacrylamide gels.

Base Sequence↗
PubMed1983

Identification of carbohydrate-binding proteins from mouse and human fibroblasts.

Three carbohydrate-binding proteins (Mr 35 000, 16 000 and 13 500) were isolated from extracts of mouse 3T3 fibroblasts by affinity chromatography on polyacrylamide beads to which was covalently bound the ligand 6-aminohexyl 4-beta-D-galactosyl-2-acetamido-2-deoxy-beta-D-glucopyranoside. None of these proteins bind to polyacrylamide beads coupled with either 6-aminohexanol or 6-aminohexyl beta-D-galactopyranoside. Therefore they appear to be carbohydrate-binding proteins specific for galactose-terminated glycoconjugates. A carbohydrate-binding protein was also purified from extracts of human foreskin fibroblasts. This protein (Mr 35000) may represent the human counterpart of the mouse protein of similar Mr and binding properties.

Animals↗
PubMed1982

Differential typing of Salmonella agona: type divergence in a new serotype.

An international collection of 419 isolates of Salmonella agona was phage typed, biotyped and colicine typed. Of 16 recognized phage types, 15 were represented. Three phage types (I, V and XVI) accounted for 84% of all isolates, were widely distributed and may be interconvertible. Biotyping afforded little type differentiation; thus 92.6% of the isolates belonged to biotype 1 a. A rhamnose non-fermenting variant line (of biotype 5a) became established in Zaire from 1979 to 1980. A maltose late-fermenting line of biotype 1 a, isolated in Scotland in 1974, did not thereafter become established. Two Col+ lines (producing colicine I b) accounted for 45 of 68 colicinogenic isolates. The implication of type diversification and the phylogenetic significance of these findings are discussed.

Amino Acids↗
PubMed1980

Interaction of ribulosebisphosphate carboxylase/oxygenase with transition-state analogues.

2-C-Carboxy-D-ribitol 1,5-bisphosphate and 2-C-carboxy-D-arabinitol 1,5-bisphosphate have been synthesized, purified, and characterized. In the presence of Mg2+, 2-C-carboxy-D-arabinitol 1,5-bisphosphate binds to ribulose-1,5-bisphosphate carboxylase/oxygenase by a two-step mechanism. The first, rapid step is similar to the binding of ribulose 1,5-bisphosphate or its structural analogues. The second step is a slower process (k = 0.04 s-1) and accounts for the tighter binding of 2-C-carboxy-D-arabinitol 1,5-bisphosphate (Kd less than or approximately to 10(-11) M) than of 2-C-carboxy-D-ribitol 1,5-bisphosphate (Kd = 1.5 X 10(6) M). Both carboxypentitol bisphosphates exhibit competitive inhibition with respect to ribulose 1,5-bisphosphate. 2-C-(Hydroxymethyl)-D-ribitol 1,5-bisphosphate and 2-C-(hydroxymethyl)-D-arabinitol 1,5-bisphosphate were also synthesized; both are competitive inhibitors with respect to ribulose 1,5-bisphosphate with Ki = 8.0 X 10(-5) M and Ki = 5.0 X 10(-6) M, respectively. Thus, the carboxyl group of 2-C-carboxy-D-arabinitol 1,5-bisphosphate is necessary for maximal interaction with the enzyme. Additionally, Mg2+ is essential for the tight binding of 2-C-carboxy-D-arabinitol 1,5-bisophsphate. A model for catalysis of ribulose 1,5-bisphosphate carboxylation is discussed which includes a functional role for Mg2+ in the stabilization of the intermediate 2-C-carboxy-3-keto-D-arabinitol 1,5-bisphosphate. Mechanistic implications that arise from the stereochemistry of this intermediate are also discussed.

Carboxy-Lyases↗
PubMed1980

Enzymatic synthesis and carbon-13 nuclear magnetic resonance conformational studies of disaccharides containing beta-D-galactopyranosyl and beta-D-[1-13C]galactopyranosyl residues.

Partially purified UDPgalactosyltransferase (EC 2.4.1.22) from bovine milk has been used to synthesize millimolar amounts of compounds such as Gal beta (1 leads to 4)Glc, Gal beta (1 leads to 4)GlcNAc-beta-hexanolamine, and Gal beta (1 leads to 4)GlcNAc beta (1 leads to 4)GlcNAc. The same method has been used to prepare similar compounds containing 13C-enriched galactopyranosyl moieties. Gal beta (1 leads to 4)GlcNAc-beta-hexanolamine was also synthesized in a solid-phase system in which the GlcNAc-beta-hexanolamine glycoside was covalently linked to agarose beads. At pH 7.0 and at 1--5 mM Mn2+ the yields of the galactosyl saccharides are greater than 90% by using 10% excess of UDPGal donor. The use of a 90% enriched [1-13C]galactosyl residue allowed the determination of the most abundant conformer about the galactopyranosyl--glycoside linkage by analysis of the carbon--carbon coupling constants from C1 of Gal to the C3', C4, and C5' of GlcNAc or Glc.

Animals↗
PubMed1980

Mass spectrometric analysis of the reactions of ribulosebisphosphate carboxylase/oxygenase.

The products of the reaction of D-[2-13C]ribulose 1,5-bisphosphate with molecular oxygen in the presence of D-ribulose 1,5-bisphosphate carboxylase/oxygenase were analyzed, after dephosphorylation, as the trimethylsilyl derivatives of glycolate and glycerate by mass spectrometry. The extent of isotopic incorporation into [1-13C]glycolate from [1-13C]glycolate 2-phosphate produced in the oxidation reaction demonstrates that at least 95% of the glycolate 2-phosphate produced arises from carbon atoms 1 and 2 of D-ribulose 1,5-bisphosphate. When D-[2-18O]ribulose 1,5-bisphosphate was used, a significant amount of [1-18O]glycolate 2-phosphate was formed, indicating that O-2 of D-ribulose 1,5-bisphosphate is retained in the carboxyl oxygens of glycolate 2-phosphate. In addition, analyses of the products of the reaction between D-[2-13C]ribulose 1,5-bisphosphate and [13C]O2 confirm and extend the conclusions of an earlier report (Müllhofer, G., and Rose, I. A. (1965) J. Biol. Chem. 240, 1341-1346) that cleavage of D-ribulose 1,5-bisphosphate occurs between carbon atoms 2 and 3 during its enzymatic carboxylation. The results eliminate possible mechanisms involving the obligatory loss of O-2 of D-ribulose 1,5-bisphosphate during its enzymatic oxidation and confirm the specificity of carbon--carbon bond cleavage in both the oxygenase and carboxylase reactions.

Carbon Isotopes↗
PubMed1980

Phage type/biotype groups of Salmonella typhimurium in Scotland 1974-6: variation during spread of epidemic clones.

Biotyping by the scheme of Duguid et al. (1975) of 2010 cultures of Salmonella typhimurium received by the Scottish Salmonella Reference Laboratory in 1974--6, the definitive phage types of which were known, revealed 137 different phage type/biotype groups. Four major epidemic clones, comprising 52% of the cultures, were recognized: 1/2a, 49/26a, 56/17g and 141/9f. The sources of each of these four groups of strains were primarily bovine (587 cultures) and human (361), suggesting a close association between infections in the two hosts. Epidemiological evidence showed that most of the outbreaks were caused by cultures of a single phage type/biotype, suggesting that both phage typing and biotyping characters were usually stable in the course of spread of epidemic strains. Thirty-two of the 63 phage types contained strains of more than one biotype. Cultures from 11 of the phage types were of two or more closely related biotypes and those from 21 others were of unrelated or distantly related biotypes. The combined use of phage typing and biotyping made it possible to detect occasional variations in the phage type or biotype in epidemic clones during their spread, e.g. phage type 49 to 204, 56 to 193, 141 to 193 and biotype 2a to 10a, 9f to 9bf, or 9cf, 26a to 26f.

Bacteriocin Plasmids↗
PubMed1979

Carbon-13-enriched carbohydrates: preparation of triose, tetrose, and pentose phosphates.

Three-, four-, and five-carbon aldononitrile phosphates were prepared, purified, and catalyticlly reduced with palladium--barium sulfate (5%) to the corresponding aldose phosphates in high yields at pH 1.7 +/- 0.1 and atmopsheric pressure. DL-Glyceraldehyde 3-phosphate and the tetrose 4-phosphates were prepared with carbon-13 enrichment at C-1, while the pentose 5-phosphates were prepared with enrichment at C-1 and C-2. Preparations of glycolaldehyde phosphate and d-glyceraldehyde 3-phosphate by lead tetra-acetate oxidation of glycerol phosphate and fructose 6-phosphate, respectively, are described. The proportions of cyclic hemiacetals and linear gem-diol forms of the two- to five-carbon aldose phosphates in aqueous solution are reported. Carbon-13 chemical shifts and carbon--phosphorus and carbon--hydrogen coupling constants for the furanose phosphate ring and linear gem-diol phosphates are reported and discussed. d-[2(-13)C]Ribulose 1,5-bisphosphate and L-[3,4(-13)C]sorbose 1,6-bisphosphate were prepared enzymatically from D-[2(-13)C]ribose 5-phosphate and dl-[1(-13)C]glyceraldehyde 3-phosphate, respectively.

Carbohydrate Epimerases↗
PubMed1979

alpha and beta-glycopyranosyl phosphates and 1.2-phosphates. Assignments of conformations in solution by 13C and 1H NMR.

The 1H and 13C NMR parameters of the anomeric pairs of aldopyranosyl phosphates and their rigid 1,2-phosphate derivatives are reported.The derivatives of D-glucose, D-galactose, and D-mannose exist in the 4C1 conformation while the L-fuco derivatives are in the C4 conformation. As judged by 31P--1H and 31P--13C coupling constants, all of the alpha anomers of the aldopyranosyl phosphates have the phosphate moiety predominantly trans to C(2) while in the beta anomers other rotamers make significant contributions. This relationship remains the same for the biologically important nucleoside diphosphate sugars (UDPGlc, UDPGal, GDPMan, and GDPFuc). From the pH dependence of 13C chemical shifts, observed in 0.5 M solutions, the pK'a2 of the alpha anomers is 6.1 while the pK'a2 of the beta anomers is 0.6--0.8 pH unit lower. In the 1.2-phosphates, the chair conformation of the parent aldose is retained while an envelope conformation is formed by the cyclic phosphate. In the alpha anomers, the plane is formed between C(2), C(1), O(1), and P while O(2) is above the plane. In the beta anomers, O(1) is out of the plane formed by the other atoms. The beta anomers have phosphorus coupled to C(3) with coupling constants of 10.8--11.7 Hz, approximately 2 Hz greater than the maximum reported for trans coupling (Lapper, R. D., & Smith, I. C. P. (1973) J. Am. Chem. Soc. 95, 2880).

Fourier Analysis↗