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Biomedical subjects

W S Hancock

Publications and source records attributed to W S Hancock.

At least 55 records · Page 3Linked to original sources
PubMed1986

Peripheral neuropathy in cats with inherited primary hyperchylomicronaemia.

Primary hyperlipoproteinaemia (hyperchylomicronaemia) with a slight increase in very low density lipoprotein) is described in 20 cats. Fasting hyperlipaemia, lipaemia retinalis and peripheral neuropathies were the most frequently detected clinical signs. The disease is thought to be inherited as an autosomal recessive trait but the exact mode of inheritance has not been determined. Affected cats showed reduced lipoprotein lipase activity measured after heparin activation compared with the response in normal cats. Plasma triglyceride and cholesterol were increased in all the cats with the major proportion of triglyceride and cholesterol being present in chylomicrons. The peripheral nerve lesions were caused by compression of nerves by lipid granulomata. It is probable that the lipid granulomata result from trauma because the nerves most often affected were at sites like the spinal foraminae where they were susceptible to trauma.

Animals↗
PubMed1985

Selective labelling of apolipoproteins A-I and C-I at methionine residues by (3H) methyl exchange.

Apolipoproteins C-I and A-I were radioactively labelled with tritium by (3H)-methyl exchange. The methionine residues were first methylated with (3H)-methyl iodide at pH 4 and the reaction products were purified by gel filtration and cation exchange chromatography. The products were then demethylated with 2-mercaptoethanol (6 M) at pH 8.6 to regenerate the apolipoproteins in an unmodified but tritiated form. The specific radioactivity for apolipoprotein C-I and A-I was 3.5 X 10(6) and 1.5 X 10(7) dpm/pmol respectively. The properties of (3H)-apolipoprotein C-I were examined by reversed phase HPLC and by incorporation into very low density lipoproteins (VLDL).

Apolipoprotein A-I↗
PubMed1985

Synthesis of peptide labelled with p-iodophenylalanine which corresponds to the first amphipathic region of apolipoprotein C-I.

The amino terminal fragment (1-15) of apolipoprotein C-1, H-Thr-Pro-Asp-Val-Ser-Ser-Ala-Leu-Asp-Lys-Leu-Lys-Glu-Phe14-Gly was prepared by the solid phase method. However, the phenylalanine residue at position 14 was replaced with p-iodophenylalanine in the chemical synthesis. The preparation of t-butyloxy-carbonyl-p-iodophenylalanine is described. After completion of the synthesis, the product was deprotected and cleaved from the resin with liquid HF. The peptide was purified by gel filtration on Sephadex G-25 and preparative high performance liquid chromatography. The purified material was shown to be homogeneous by amino acid analytical data and by chromatography in three different analytical reversed phase HPLC systems. The peptide was then labelled by the chloroamine-T procedure and good incorporation of 125I was obtained. After purification of the product by gel filtration on Biogel P-2, the labelled pentadecapeptide was tested for ability to bind to Very Low Density Lipoproteins (VLDL) in the following manner: VLDL was isolated from normolipemic serum by ultracentrifugal flotation and 100-microliter samples were incubated with labelled material dissolved in 200 microliter of 0.5 M NaCl, 0.02 M sodium phosphate buffer, pH 7.4 at 37 degrees for 30 min. The VLDL fraction was again isolated by ultracentrifugal flotation and the incorporation of radioactivity into the lipoprotein was measured. Under these conditions, a sample of [3H]-native apolipoprotein C-I was incorporated to an extent of 83% of the total sample, while the [125I]-pentadecapeptide exhibited an incorporation of 8.7%.

Amino Acid Sequence↗
PubMed1985

Rotavirus neutralizing protein VP7: antigenic determinants investigated by sequence analysis and peptide synthesis.

The rotavirus neutralizing antigen, VP7, is a 37,000-molecular-weight glycoprotein which is a major component of the outer shell of the virion. The amino acid sequence of VP7 for strain S2 (human serotype 2) and Nebraska calf diarrhea virus (bovine serotype) has been inferred from the nucleic acid sequence of cloned copies of genomic segment nine. Comparison of the amino acid sequences of these two VP7 proteins with those already determined for other rotavirus strains reveals extensive sequence conservation between serotypes with clusters of amino acid differences sited predominantly in hydrophilic domains of the protein. Six peptides have been synthesized that span the hydrophilic regions of the molecule. Antisera to these peptides both recognize the respective homologous peptides in a solid-phase radioimmunoassay and bind to denatured VP7 in a Western blot. However, none of the antisera either recognize virus or exhibit significant neutralizing activity, indicating that these peptide sequences are not available on the surface of the virus.

Amino Acid Sequence↗
PubMed1984

Antisera to synthetic peptide recognize high molecular weight enkephalin-containing proteins.

Antisera to a synthetic peptide corresponding to the 95-117 sequence of proenkephalin were used to develop a sensitive radioimmunoassay. Gel-filtration of acid extracts of bovine adrenal medulla and purified chromaffin granules revealed that the antisera recognized high molecular weight material (Mr approximately 5,000-30,000). The material in peak I ( Mr 20 ,000-30,000) and peak II (Mr 10,000-20,000) was further purified by immunoaffinity chromatography. Sequential digestion of each of these fractions with trypsin and carboxypeptidase B generated immunoreactive Met-enkephalin. This study demonstrates that antisera against a synthetic peptide cross-react with high molecular weight enkephalin-containing precursors, validating the use of these antisera in studies of enkephalin biosynthesis.

Adrenal Medulla↗
PubMed1983

Occurrence of idiopathic, familial hyperchylomicronaemia in a cat.

Primary hyperlipoproteinaemia (hyperchylomicronaemia with slight very low density lipoprotein elevation) is described in two related male cats. Fasting hyperlipaemia, lipaemia retinalis and subcutaneous xanthomas were detected on clinical examination. In one cat lipoprotein lipase activity measured after heparin activation was significantly reduced compared to the response in a normal cat. The lipid and protein concentration in each of the lipoprotein classes and the lipoprotein distribution of the two hyperlipaemic cats, two normolipaemic relations and 16 normolipaemic adult cats were determined. Plasma cholesterol and triglyceride levels were elevated in the hyperlipaemic cats with the major proportion of triglyceride and cholesterol being present in chylomicrons whereas in normolipaemic cats the majority of triglyceride was contained in very low density lipoprotein. High density lipoprotein was the predominant lipid carrier in both the normolipaemic and the hyperlipaemic cats but the protein content in chylomicrons was elevated in the two affected cats. The lipoprotein distribution in normal cats in this study agrees with previously reported values. The hyperlipaemic cats showed many of the features of familial lipoprotein lipase deficiency (type I hyperlipoproteinaemia, exogenous chylomicronaemia) which is an inherited disease in man.

Animals↗
PubMed1983

Hyperbetalipoproteinaemia and hyperalphalipoproteinaemia in a single family.

Twelve members of a kindred were studied of whom six showed elevated low-density lipoprotein (LDL) cholesterol levels. Within the blood related group nine showed elevated high density lipoprotein (HDL) cholesterol levels and correspondingly elevated HDL lipoprotein apoprotein levels. The members with elevated LDL also therefore had elevated HDL. There was no family history of premature vascular disease in the kindred and the 74 year old member was clinically free of ischaemic vascular disease. It is considered that the coincidence of a familial tendency to high blood LDL and HDL was not associated with appearance of premature arterial disease in this kindred.

Adolescent↗
PubMed1981

Application of 1,1'-carbonyldiimidazole-activated matrices for the purification of proteins. III. The use of 1,1'-carbonyldiimidazole-activated agaroses in the biospecific affinity chromatographic isolation of serum antibodies.

The characteristics of ligands immobilised into 1,1'-carbonyldiimidazole-treated cross-linked agarose have been further evaluated. Since the intermediate activated matrix (an imidazolyl carbamate-agarose) is susceptible to nucleophilic attack by free amino groups, but is relatively stable to oxygen nucleophiles, ligands ranging from simple organic primary amines, amino acids, proteins and other biological substances, which contain amino group functionality, can be bound to the agarose via a N-alkylcarbamate (urethane) linkage. This covalent linkage has been found to exhibit good chemical stability to mildly acidic and basic elution conditions. The use of 1,1'-carbonyldiimidazole-activated agarose in the biospecific affinity chromatography of immunoglobulins, present in normal and pathological sera, is described.

Antibodies↗
PubMed1981

The use of reversed-phase high-performance liquid chromatography with radial compression for the analysis of peptide and protein mixtures.

This report describes the separation of peptide and protein mixtures on a C18- microparticulate support which was packed in a polyethylene cartridge and subjected to radial compression of ca. 2600 p.s.i. The purpose of the radial compression was to minimise inhomogeneities in the column packing, in particular in the region of the column wall and end fittings. The effectiveness of this new chromatographic system was demonstrated by the efficient separation of the following mixtures: the C-apolipoproteins isolated from human very low density lipoproteins; the polymorphs of apolipoprotein A-I; the tryptic fragments of apolipoprotein C-II; the complex mixture generated by partial proteolysis of apolipoprotein B; the tryptic fragments of 3H- and 14C-labelled beta-chain of murine IA alloantigen and the tryptic fragments of carboxymethylated lambda chain isolated from human immunoglobulin G. The separated peaks were identified by amino acid analysis, radioactivity counting and in the last example by amino acid sequence determination. The mobile phase consisted of 1% aqueous solution of triethylammonium phosphate, pH 3.2 with acetonitrile or isopropanol as the organic modifier.

Amino Acids↗
PubMed1981

Use of mixed-mode, high-performance liquid chromatography for the separation of peptide and protein mixtures.

The packing material recently introduced for use in a radial compression chamber, with a partially flexible cartridge, has a low C18-coating (5% w/s) combined with an absence of secondary capping. This report demonstrates that such a support, which contains significant concentrations of both free silanol and hydrocarbon groups, can allow a mixed-mode separation to occur via adsorption and reversed-phase separation mechanisms. In any given separation, the predominant mechanism depends both on the nature of the sample and the mobile phase. For efficient peptide and protein separations, it was necessary to suppress most silanol group interactions by the use of a mobile phase which contained a high concentration of an amine phosphate, e.g., 0.17 M triethylammonium phosphate, pH 3.2. In addition, it was necessary to deactivate further the silanol groups by an initial column wash of at least 20 column volumes of methanol. Samples which contained strongly basic groups, for example the guanidino group of arginine, can still exhibit poor separation efficiencies on such a support. These problems were largely overcome, however, with the use of isopropanol as an organic modifier. If these precautions were followed, the packing material gave excellent selectivities in the separation of closely related materials, as well as allowing increased sample capacities. These observations will be supported by an examination of the chromatographic properties of a range of small peptides, the C-apolipoprotein mixture present in human very-low-density lipoproteins and the purification of an 8-mg sample of a synthetic pentadecapeptide in a single chromatographic run.

Apolipoproteins↗
PubMed1981

Separation of the apoprotein components of human very low density lipoproteins by ion-paired, reversed-phase high performance liquid chromatography.

A number of crude apolipoprotein samples isolated from human very low density lipoproteins (VLDL) were analyzed by reversed phase high performance liquid chromatography. The mobile phase consisted of a 1% solution of the polar ion-pairing reagent triethylammonium phosphate. A slow, nonlinear gradient of acetonitrile (37--42%) was used to elute the apolipoproteins. The order of elution was as follows: apolipoprotein CX, apolipoprotein C-I, apolipoprotein C-III2, apolipoprotein C-III1, apolipoprotein C-IIIQ and apolipoprotein C-II. This order is consistent with the known polarity of the proteins, i.e., the most nonpolar, apolipoprotein C-II, was the last to be eluted, whereas apolipoprotein C-I, with the lowest nonpolar surface area eluted first. The recovery of the individual apolipoproteins was 80--95% and the individual peaks were characterized by amino acid analysis, UV absorption spectra amd chromatography of pure protein standards.

Acetonitriles↗
PubMed1981

Use of the chromogenic p-(p-(dimethylamino)phenylazo) benzyl (AZ) ester in the synthesis of leu-enkephalin.

Solution synthesis of Leu-enkephalin has been achieved using the coloured carboxyl protecting p-(p-(dimethylamino)phenylazo)benzyl moiety throughout the synthesis. The intense colour of this protecting group allowed facile purification of protected intermediates on silica gel or ion exchange columns and visualisation on thin-layer chromatography. The chromogenic p-(p-(dimethylamino)phenylazo)benzyl ester group readily withstands successive treatments with 25% trifluoroacetic acid in dichloromethane yet is cleanly removed by catalytic hydrogenation.

Amino Acids↗
PubMed1981

Use of perfluoroalkanoic acids as volatile ion pairing reagents in preparative HPLC.

This report describes the successful use of the ether soluble, ion pairing reagents perfluoropropoinic and perfluorobutyric acid in the preparative and analytical reserved phase HPLC of underivatised peptides. The preparative separation of a 1-g sample of Pyr-His-Gly, the proposed anorexigenic peptide, is described on C18-silica which was packed in a flexible-walled cartridge and subjected to radial compression. The mobile phase consisted of an aqueous solution of perfluorobutyric acid (5 mM) and a flow rate of 100 ml/min was used. The purified peptide was simply isolated by neutralizing and freeze-drying the corresponding peak and then extracting the excess ion pairing reagent with ether. The product was then shown to be homogeneous by analytical HPLC and amino acid analysis. The tripeptide failed to show any effect on food intake, water intake or body weight in female rats. Similarly no effect was noted on the reproductive cycles of the rat.

Animals↗