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Biomedical subjects

N M Young

Publications and source records attributed to N M Young.

At least 73 records · Page 4Linked to original sources
PubMed1988

The solution structure of concanavalin A probed by FT-IR spectroscopy.

The secondary structural properties of various forms of concanavalin A in solution were investigated by Fourier-transform infrared spectroscopy in the Amide I region. As in the crystal, the solution structure of the native protein consists mainly of antiparallel beta-sheet. Carbohydrate binding does not produce major changes in the overall secondary structure of concanavalin A, but affects infrared bands due to loops and beta-turns. Upon demetallization, the spectrum of concanavalin A shows only a small change in the Amide I band, indicating that whereas the beta-sheet structure is conserved, the tertiary properties may be altered. There are also changes in the bands from the tyrosine residues which are compatible with local changes in structure. Confirming tertiary structural differences, the cation-depleted apoprotein is much less stable, denaturing around 63 degrees C, while the native protein denatures only at temperatures around 85 degrees C. Tetramerization proceeds without significant secondary structural change. However, aggregation of the tetramers leads to a significant decrease of the bands corresponding to beta-sheet structure, and changes in the tyrosine bands.

Calcium↗
PubMed1988

Surgical treatment of vertigo with retrolabyrinthine vestibular neurectomy.

Results for control of vertigo and preservation of hearing in patients who have had a retrolabyrinthine vestibular neurectomy (RVN) by our group were analyzed retrospectively. This procedure consists of selective section of the vestibular nerve in the posterior cranial fossa. Vertigo was completely controlled in all but two of 31 patients, one of whom required revision surgery to control attacks. Analysis of these two cases suggests that the cause of persistent vertigo is incomplete neurectomy. With our current surgical technique in patients with Meniere's disease, hearing results were not statistically different from our results with surgery of the endolymphatic sac. Control of vertigo was much more successful with the RVN than endolymphatic sac surgery.

Adult↗
PubMed1987

The carbohydrate specificity of conglutinin and its homology to proteins in the hepatic lectin family.

Inhibition experiments with D-mannose oligosaccharides establish that conglutinin recognises terminal alpha 1----2 mannobiosyl units present in the glycopeptide of the alpha-chain of the complement component C3b. On the basis of its three domain structure and the homology of its N-terminal amino-acid sequence to that of the dog pulmonary surfactant protein, it is proposed that conglutinin is a member of the hepatic lectin family.

Animals↗
PubMed1987

The dinitrophenyl group as a selective label in high-performance liquid chromatography of peptides.

1-Fluoro-2,4-dinitrobenzene can be used to selectively label histidine, tyrosine, and cysteine residues in maleylated proteins. The usefulness of the resulting chromophores for peptide mapping by high-performance liquid chromatography was demonstrated with the lectin from sainfoin (Onobrychis viciifolia). The 2,4-dinitrophenyl (Dnp) label also can be used in a hydrophobic modulation approach as the mobility of a labeled model peptide changes considerably when its Dnp group is removed by thiolysis. Application of the method for checking sequences obtained by DNA or amino acid methods was shown by experiments with Viciae lectins. The probable cleavage site that generates the pea lectin's beta-chain from the alpha-beta precursor was identified and the sequence differences between the lentil and pea lectin beta-chains were examined.

Chromatography, High Pressure Liquid↗
PubMed1987

Studies on alternaria allergens. VI. Stability of the allergen components of Alternaria tenuis extracts under a variety of storage conditions.

The allergenic potencies of samples of two commercial Alternaria tenuis extracts stored under various conditions were compared at 3 months intervals over a period of 1 year. Aliquots of the extracts at 5.12 mg/ml were maintained in: saline; 50% glycerol-saline mixture; 0.4% phenol-saline; 0.4% phenol-saline containing 0.03% human serum albumin, and samples were also kept in lyophilized form. The samples were stored at -20, 4, 24, 37 and 56 degrees C. Their potency was measured by radioallergosorbent inhibition assay and by mouse IgE passive cutaneous anaphylaxis (PCA) tests. The lyophilized extracts fully maintained their activity at all temperatures for 12 months. Aqueous extracts with or without preservatives also maintained their potency if stored at -20, 4 and 24 degrees C, but showed a significant loss of PCA activity at 56 degrees C, after 3 months. A deleterious effect of phenol was observed in one of the extracts stored at -20 degrees C for 3 months, by PCA tests. The use of preservatives to retain potency over a period of 12 months is therefore unnecessary and may be deleterious. The results indicate that, while storage of A. tenuis in a lyophilized form is best at all temperatures, aqueous extracts are also remarkedly stable when stored at -20, 4 and 24 degrees C, over a 12-month period.

Allergens↗
PubMed1986

Determination of chemical properties of individual histidine and tyrosine residues of concanavalin A by competitive labeling with 1-fluoro-2,4-dinitrobenzene.

A selective peptide-mapping procedure was devised to purify peptides containing histidine or tyrosine residues from proteolytic digests of concanavalin A (Con A). The protein was modified with maleic anhydride followed by 1-fluoro-2,4-dinitrobenzene (Dnp-F) and then digested with thermolysin. The resulting labeled peptides were separated by high-performance liquid chromatography, and the Dnp-histidine and Dnp-tyrosine peptides were identified by their spectral characteristics. From their amino acid compositions, the labeled peptides could all be assigned within the known sequence. Peptides representing five of the six histidines and all seven tyrosines were obtained. With the same peptide-mapping procedure, the chemical properties (pK and reactivity) of these residues were determined. Samples of concanavalin A at various pH values were labeled with trace amounts of [3H]Dnp-F, in the presence of Gln-Gly as an internal standard. To each sample was added an aliquot of a mixture of [14C]Dnp-Gln-Gly and [14C]Dnp-maleyl-Con A. Portions of each sample were removed, [14C]Dnp-Ala-Ala and epsilon-[14C]Dnp-lysine were added, and the mixtures were hydrolyzed. The various Dnp amino acid derivatives were purified by HPLC. The remainder of each [3H]Dnp sample was maleylated, dinitrophenylated, and digested with thermolysin and separated by HPLC as above. From the 3H/14C ratios of the Dnp amino acid derivatives and the Dnp peptides relative to the ratio of the internal standard, pK and reactivity data were obtained for (a) the average behavior of the lysine, histidine, and tyrosine residues and (b) the individual behavior of the N-terminal alanine residue and the five histidine and seven tyrosine residues in the protein.(ABSTRACT TRUNCATED AT 250 WORDS)

Amino Acid Sequence↗
PubMed1985

The thiol groups of mouse immunoglobulin A. Incomplete formation of the C alpha 1-domain disulphide bridge.

The BALB/c IgA (immunoglobulin A) myeloma protein M167 contained on average 5.7 free SH groups per IgA dimer. These groups were preponderantly on the heavy chains and comprised two distinct populations: 3.3 exposed SH groups per dimer in the Fc region, and 2.4 buried SH groups per dimer in the Fd region, detectable o only after denaturation. To locate the cysteine residues involved, labelled peptides were purified from thermolysin digests of radioalkylated IgA by high-performance liquid chromatography. From the amino acid compositions of the peptides, the exposed thiol groups were assigned to Cys-307 in the C alpha 2 domain, which thus existed in the reduced form to an extent exceeding 80%. This residue may allow attachment of secretory component to dimer IgA in the mouse to proceed via thiol-disulphide exchange. The buried thiol groups were assigned to Cys-150 and Cys-208, in the C alpha 1 domain, each being in the reduced form to the extent of approx. 30%. This pair of residues would normally give rise to the characteristic intradomain disulphide bridge. It appears that disulphide formation is not a crucial event during folding of the C alpha 1 domain in IgA biosynthesis. The sequence in the region 140-151 was re-investigated, and residue 142 was shown to be serine, not cysteine, helping explain the lack of heavy-chain-light chain bonding in BALB/c mouse IgA. A disulphide-bond model for mouse IgA is proposed on the basis of these assignments and other features of the mouse alpha-chain sequence.

Animals↗
PubMed1985

The circular dichroism of phosphocholine-specific mouse hybridoma and myeloma proteins: unusual properties of the hybridoma protein 101.6G6.

The circular dichroism (CD) spectra of five myeloma and six hybridoma proteins specific for phosphocholine were measured in the 250-310-nm range. The effect on the CD spectra of adding phosphocholine was also examined. The five myeloma proteins all had distinctive native spectra and, except for M603 and W3207, unique changes occurred on ligand binding. The hybridomas were chosen as pairs from each of the three known families of phosphocholine-specific immunoglobulins. Those from the T15 or M603 families resembled the appropriate prototype. However, the proteins from the M167 family were all distinctively different in their CD properties. In particular, the hybridoma protein 101.6G6 showed large CD changes on hapten binding and values for the association constant for phosphocholine of 1.1 X 10(5) M-1 and of 5.8 X 10(2) M-1 for acetylcholine were obtained by CD spectrophotometric titration. The CD properties of the proteins are interpreted in the light of the sequence data so far available, including the possible role of the D-segment.

Acetylcholine↗
PubMed1985

Assignment of lectins specific for D-galactose or N-acetyl-D-galactosamine to two groups, based on their circular dichroism.

The circular dichroism (CD) spectra of thirteen lectins, most being specific for D-galactose or N-acetyl-D-galactosamine, were compared. Two groupings are proposed on the basis of the CD in the near-ultraviolet region. Group one comprises the lectins from Arachis hypogaea, Glycine max, Phaseolus coccineus, P. lunatus, P. vulgaris, and Vicia villosa, and group two comprises lectins from Caragana arborescens, Cytisus sessilifolius, Dolichos biflorus, Griffonia simplicifolia, and Wisteria floribunda. The CD spectra of lectins from Bauhinia purpurea and Sophora japonica were different from any of the other lectins. CD difference spectra produced by the two sugars were distinctive for each protein, suggesting that the combining sites of these lectins are not homologous despite similarities in specificity and that the CD spectral similarities arise from residues in other, more homologous regions of the proteins.

Acetylgalactosamine↗
PubMed1985

Studies on Alternaria allergens. V. Comparative biochemical and immunological studies of three isolates of Alternaria tenuis cultured on synthetic media.

Six extracts were prepared from Alternaria tenuis isolated ATCC 6663, ATCC 16086 and DAOM (Agriculture Canada) 183905 grown separately on two synthetic media, revised tobacco and Czapek's, and their biochemical and immunological properties were examined. High-performance liquid chromatography revealed considerable variations in UV absorbance and carbohydrate profiles among extracts from the different isolates. These differences were less marked among samples of the same isolate cultured on different media. Enzyme screening showed that all extracts contained large amounts of phosphatases and glucosidases and moderate quantities of esterases. Only the alpha-galactosidase activity showed any correlation with allergenic activity. No significant variation was observed in isoelectric focusing patterns. Extensive antigenic cross-reactivity even between the different isolates was found in precipitin studies. In mouse IgE passive cutaneous anaphylaxis tests, all extracts gave reactions of similar intensity. In direct RAST and RAST inhibition assays, ATCC 16086 grown on revised tobacco medium was found to be the most potent and approached the activity of an extract from a commercial material (B-I). DAOM 183905 grown on either medium was next in potency while ATCC 6663 samples were the least potent. The results indicate that it is possible to obtain extracts of high allergenic potency for standardization purposes from growth of selected A. tenuis isolates on a chemically defined medium.

Allergens↗
PubMed1984

Structural differences between two lectins from Cytisus scoparius, both specific for D-galactose and N-acetyl-D-galactosamine.

Three lectin fractions were obtained from seeds of the leguminous plant Cytisus scoparius (Scotch broom) by means of affinity chromatography on a N-acetyl-D-galactosamine medium. The first fraction, termed CSIa, was equally well inhibited in haemagglutination experiments by D-galactose and by N-acetyl-D-galactosamine and consisted of a group of isolectins formed from closely related polypeptide chains of approx. Mr 30000. The second fraction, CSIb, was closely related to CSIa in specificity, c.d. and other properties. The third fraction contained a homogeneous lectin, CSII, formed from subunits again of approx. Mr 30000. CSII was 100 times more readily inhibited by N-acetyl-D-galactosamine than by D-galactose. Despite the similarity in specificity, comparative studies of their amino acid composition, c.d. and N-terminal amino acid sequence showed that the CSIa and CSII lectins diverged considerably in structure. The lectin from Cytisus sessilifolius, specific for chitobiose, was also examined and resembled CSIa in composition and c.d. properties.

Acetylgalactosamine↗
PubMed1984

Studies on alternaria allergens. IV. Comparative biochemical and immunological studies of commercial Alternaria tenuis Batches.

The biochemical and immunological characteristics of a series of commercial samples of Alternaria tenuis were examined, including samples from two different companies. There was a considerable variation in the biochemical parameters such as the gel filtration profile on Sephadex G-100, isoelectric focussing pattern and immunodiffusion precipitin band pattern. The content and proportion of two high molecular weight polysaccharides, one a galactomannan and the other a glucan, also varied greatly. In contrast, the allergenic properties of the samples were more consistent. In rat IgE passive cutaneous anaphylaxis tests, extensive cross-reactivity was observed among batches from the same company, but this cross-reactivity was weaker when batches from the two companies were compared. In direct radioallergosorbent tests (RAST) the samples showed similar reactivities, but RAST inhibition tests showed a 40-fold range in activity between samples from one company and 55-fold between batches from the two sources. It was found that the proportions of the major A. tenuis allergen and a related hypoallergenic antigen in the samples were relatively constant, hence RAST inhibition can be used as a measure of allergenic potency despite the presence of this cross-reacting antigen. The lack of correlation between allergenic and biochemical properties of the various samples suggest that a simple biochemical test is unlikely to be a satisfactory replacement for the RAST inhibition assay in standardizing A. tenuis extracts.

Allergens↗
PubMed1983

Magnesium as a natural substitute for manganese in concanavalin A and other lectins.

Addition of magnesium to apo-concanavalin A in the presence of calcium was shown by ultraviolet difference spectroscopy to generate a holoprotein spectroscopically identical to the MnCa-holoprotein. The MgCa- and MnCa-forms bound equally strongly to Sephadex G-75. In kinetic experiments, the binding of Mg2+ was much slower than Mn2+ binding; Kd for Mg2+ was estimated as 7.4 mM. The combined Mg2+ and Mn2+ contents of 10 lectins specific for D-galactose or N-acetyl-D-galactosamine were each close to one atom per subunit, suggesting occupancy of the Mn2+ site by Mg2+ is common in plant lectins.

Chemical Phenomena↗
PubMed1983

Conformational heterogeneity of the copper binding site in azurin. A time-resolved fluorescence study.

Comparison of the fluorescence spectra and the effect of temperature on the quantum yields of fluorescence of Azurin (from Pseudomonas fluorescens ATCC-13525-2) and 3-methylindole (in methylcyclohexane solution) provides substantive evidence that the tryptophan residue in azurin is completely inaccessible to solvent molecules. The quantum yields of azurin (CuII), azurin (CuI), and apoazurin (lambda ex = 291 nm) were 0.052, 0.054, and 0.31, respectively. Other evidence indicates that there is no energy transfer from tyrosine to tryptophan in any of these proteins. The fluorescence decay behavior of each of the azurin samples was found to be invariant with emission wavelength. The fluorescences of azurin (CuII) and azurin (CuI) decay with dual exponential kinetics (tau 1 = 4.80 ns, tau 2 = 0.18 ns) while that of apoazurin obeys single exponential decay kinetics (tau = 4.90). The ratio of pre-exponentials of azurin (CuII), alpha 1/alpha 2, is found to be 0.25, and this ratio increases to 0.36 on reduction to azurin (CuI). The results are interpreted as originating from different interactions of the tryptophan with two conformers of the copper-ligand complex in azurin.

Azurin↗
PubMed1983

The circular dichroism of ovoverdin and other carotenoproteins from the lobster Homarus americanus.

The circular dichroic (CD) spectra of the alpha-, beta'-, and gamma-crustacyanins, ovoverdin, and the yellow lobster-shell protein were measured in the region 200-750 nm, for comparison with the CD spectrum of the free carotenoid astaxanthin. The two carotenoid chromophores of ovoverdin gave a CD spectrum with a series of bands of alternating sign and ellipticities up to 1.9 X 10(5) degree X cm2 X dmol-1, comparable to the low temperature CD spectrum of astaxanthin in the UV region. The visible region of ovoverdin also contained strong CD bands where astaxanthin itself has very weak ones. The blue (640 nm) chromophore of ovoverdin gave a broad negative CD feature quite different from the blue chromophores of the three crustacyanins. The crustacyanins have a broad positive feature between 400 and 610 nm with ellipticities up to 2 X 10(5) degree X cm2 X dmol-1, followed at higher wavelengths by a negative band with an ellipticity up to 1 X 10(5) degree X cm2 X dmol-1. Gaussian curve-fitting procedures showed the positive features to consist of a minimum of two or three CD bands. In addition to the exciton bands at the main visible absorption, the yellow lobster-shell protein had a pair of CD bands of equal but opposite ellipticity associated with an absorption band between 250 and 340 nm. The UV regions of the CD spectra of the five carotenoproteins have bands from both carotenoid and protein chromophores, and possible assignments of these bands to one or the other of the two types of chromophore are proposed.

Animals↗
PubMed1982

Binding of disaccharides by peanut agglutinin as studied by ultraviolet difference spectroscopy.

The binding of the disaccharides methyl beta-D-lactoside and 2-acetamido-2-deoxy-3-O-(beta-D-galactopyranosyl)-beta-D-galactopyranose [beta-D-Gal-(l leads to 3)-D-GalNAc] to peanut agglutinin was studied by ultraviolet difference spectroscopy. The magnitude of the difference spectra varied with the concentration of the carbohydrates; association constants and thermodynamic parameters were determined from titration experiments at different temperatures. The enthalpy and entropy changes for binding of methyl beta-D-lactoside were found to be delta H degree = -65 +/- 4 kJ mol-1, delta S degree = -156 +/- 14 J mol-1 K-1. For beta-D-Gal-(1 leads to 3)-D-GalNAc the observed thermodynamic parameters were delta H degree = -78 +/- 5 kJ mol-1,, delta S degree = -177 +/- 16 J mol-1 K-1. For both disaccharides, the enthalpy change upon binding to the lectin is much larger than found for the binding site on peanut agglutinin. The observed parameters are compared with those found for the binding of monosaccharides and oligosaccharides to other lectins and to lysozyme. Molecular models of the minimum energy conformers of beta-D-Gal(1 leads to 3)-D-GalNAc and methyl beta-D-lactoside are used to interpret the interaction of these, and structurally related ligands, with the peanut agglutinin binding site.

Antigens, Tumor-Associated, Carbohydrate↗