Search PubMed⌕ Search

Biomedical subjects

L A Hunt

Publications and source records attributed to L A Hunt.

At least 37 records · Page 2Linked to original sources
PubMed1984

Nicotine-stimulated proteins in mouse cells are distinct from heat-shock proteins.

Treatment of mouse tissue-culture cells with nicotine concentrations of 1 mM or less had no significant effects on cell viability, morphology or protein synthesis, but higher concentrations resulted in both altered cell morphology (rounding and vacuolization) and alterations in [3H]leucine-labelled protein profiles on sodium dodecyl sulphate/polyacrylamide gels. The synthesis of a Mr-70 000 protein was increased more than 2-fold relative to that of other major cellular proteins in 3T3 and L929 cells treated with 5 mM-nicotine and in B16 cells treated with 10 mM-nicotine, and this protein appeared to be a soluble cytoplasmic polypeptide. The radiolabelling of several additional polypeptides (Mr 62 000 in 3T3 cells, and Mr 45 000 and 38 000 in B16 cells) was also stimulated by nicotine. The nicotine-enhanced Mr-70 000 protein was distinct, however, from a major cell stress/heat-shock protein whose synthesis was stimulated after incubation of cells at 43.5 degrees C for 20 min.

Animals↗
PubMed1983

Unusual heterogeneity in the glycosylation of the G protein of the hazelhurst strain of vesicular stomatitis virus.

The asparagine-linked oligosaccharides of the G protein of the Hazelhurst subtype of the New Jersey serotype of vesicular stomatitis virus (VSV) have been compared with the oligosaccharides from the G protein of the well-characterized Indiana serotype of VSV, with baby hamster kidney cells in monolayer culture as the host for both viruses. [3H]Glucosamine- and [3H]mannose-labeled glycopeptides from the G protein of purified virus were analyzed by the combined techniques of endo-beta-N-acetylglucosaminidase H (ENDO-H) digestion, concanavalin A and lentil lectin affinity chromatography, and Bio-Gel P-4 chromatography. Although almost all of the Indiana G protein oligosaccharides were acidic-type structures, as expected from previous studies; the Hazelhurst G protein contained a mixture of acidic-type, hybrid-type containing sialic acid, and neutral-type (predominantly Man5-6GlcNAc2-Asn) structures. The vast majority of acidic-type oligosaccharides from both the Hazelhurst and Indiana G proteins were diantennary structures, with less than half containing fucose linked to the innermost N-acetylglucosamine. Additional analysis of the Hazelhurst G protein by ENDO-H digestion and gel electrophoresis suggested that some of the mature G polypeptides contained acidic-type structures at both glycosylation sites, whereas the remainder contained an ENDO-H-resistant, acidic-type structure at one site and an ENDO-H-sensitive, hybrid- or neutral-type structure at the other site.

Animals↗
PubMed1983

Glycosidase analysis of large acidic-type glycopeptides from viral and cellular membrane glycoproteins. Evidence for a common oligomannosyl core with branch sugar heterogeneity.

The asparagine-linked oligosaccharides of the complex acidic-type from [3H]mannose-, [3H]glucosamine- or [3H]galactose-labelled membrane glycoproteins of BHK21 cells and Rous-sarcoma virus were analysed by gel filtration combined with extensive digestion with endo- and exo-glycosidases from bacterial and eukaryotic sources. The neutral products from the digestion with a mixture of exoglycosidases and endo-beta-N-acetylglucosaminidase D from Diplococcus pneumoniae included a series of [3H]mannose- and [3H]glucosamine-labelled neutral oligosaccharides that were all converted by digestion with eukaryotic beta-N-acetylglucosaminidases into free N-acetylglucosamine and a small oligomannosyl core containing two alpha-linked mannose residues and a third mannose residue beta-linked to N-acetylglucosamine. These studies suggested that the complex acidic-type oligosaccharides from cellular and viral membrane glycoproteins contained a common oligomannosyl core region (Man2 alpha leads to Man beta leads to GlcNAc2), with heterogeneity in the number and/or linkage of outer branch N-acetylglucosamine residues resulting in partial resistance to beta-N-acetylglucosaminidase from a bacterial source.

Animals↗
PubMed1983

Unusual neutral oligosaccharides in mature Sindbis virus glycoproteins are synthesized from truncated precursor oligosaccharides in Chinese hamster ovary cells.

We have previously demonstrated the presence of unusual small asparaginyl-oligosaccharides [(Man)3GlcNAc2-ASN] in the mature glycoproteins of Sindbis virus released from both wild-type and lectin-resistant Chinese hamster ovary cells, but the mechanism of synthesis of these structures was not determined. Gel filtration and endo-beta-N-acetylglucosaminidase analyses of Pronase-digested glycopeptides from [3H]mannose-labelled Sindbis virus released at different times after infection of a phytohaemagglutinin-resistant line of Chinese hamster ovary cells demonstrated that these small asparaginyl-oligosaccharides were present in similar relative amounts in virus released throughout the virus infection, rather than arising primarily at late times when cytopathic effects were maximal. Similar analyses of pulse-labelled, cell-associated viral glycopeptides suggested that these small oligosaccharides on mature virus glycoprotein resulted from the normal alpha 1,2-mannosidase processing of truncated precursor oligosaccharides (containing five rather than nine mannoses), rather than from aberrant processing or degradation of the full-size precursor oligosaccharides or normal intermediates.

Animals↗
PubMed1983

Comparison of the oligosaccharide moieties of the major envelope glycoproteins of the subgroup A and subgroup B avian myeloblastosis-associated viruses.

The nature of the oligosaccharide chains of the major envelope glycoprotein, gp85, from avian myeloblastosis-associated viruses has been examined for the subgroup A and subgroup B viruses replicated in fibroblasts from the same chicken embryos. Pronase-digested glycopeptides from [3H]mannose- or [3H]glucosamine-labeled viruses were analyzed by the combined techniques of gel filtration, endo-beta-N-acetylglucosaminidase digestion, and concanavalin A affinity chromatography. The gp85 protein from these two viruses, and also from another subgroup A avian leukosis virus replicated in the same cells, contained a diverse array of asparagine-linked oligosaccharides of the acidic type [(sialic acid +/- galactose-N-acetylglucosamine)2-4-(mannose)3-N-acetylglucosamine2(+/- fucose)-asparagine], hybrid type (sialic acid +/- galactose-N-acetylglucosamine-(mannose)5,4-N-acetylglucosamine2-asparagine), and neutral type [(mannose)5-9-N-acetylglucosamine2-asparagine], with the more highly branched (tri or tetraantennary or both) acidic-type structures representing the predominant class of oligosaccharide. Minor differences were observed between the gp85 of the subgroup B versus subgroup A viruses.

Avian Leukosis Virus↗
PubMed1982

Lectin affinity chromatography of glycopeptides and oligosaccharides from normal and lectin-resistant Chinese-hamster ovary cells.

The [3H]mannose-labelled glycopeptides from two lectin-resistant lines of Chinese-hamster ovary cells were fractionated by chromatography on lentil lectin-Sepharose and concanavalin A-agarose columns and subsequently analysed by gel filtration in comparison with the glycopeptides of the parental cell line. Essentially all of the [3H]mannose-labelled asparaginyl-oligosaccharides from the 'single-mutant' cells selected for resistance to phytohaemagglutinin and the 'double-mutant' cells selected for additional resistance to concanavalin A were not bound to lentil lectin, whereas approximately one-sixth of the parental-cell glycopeptides were bound and specifically eluted with alpha-methyl mannoside. These bound and eluted glycopeptides represented a specific subset of the complex acidic-type asparaginyl-oligosaccharides. The percentage of radiolabelled glycopeptides and oligosaccharides from each cell line that were specifically bound to concanavalin A was consistent with the relative sensitivities of the three cell lines to this lectin. The major radiolabelled species in the endoglycosidase digest of the 'double-mutant'-cell glycopeptides (Man4GlcNAc1-size neutral oligosaccharides) were not bound to concanavalin A, whereas essentially all of the other neutral-type oligosaccharides were bound. In addition, the larger neutral-type oligosaccharides (Man8--9GlcNAc1) were more strongly bound to concanavalin A than were either the smaller neutral-type or the di-antennary acidic-type structures.

Animals↗
PubMed1982

Lectin affinity chromatography of Sindbis and Rous sarcoma virus glycopeptides and oligosaccharides.

Glycopeptides and endogly cosidase-digested oligosaccharides from [3H]mannose-labeled Rous sarcoma virus and Sinbis virus have been fractionated by lentil lectin-Sepharose and concanavalin A-agarose affinity chromatography and subsequently analyzed by BioGel P-4 gel filtration. Only a specific subset of the Con A-bound asparaginly-oligosaccharides from he two viruses was also bound to lentil lectin, and this freaction apparently represented fucose-containing, diantennary acidic-type structures ((NeuNAc +/- Gal-GlcNAc)2 Man3 -GlcNAc2 (fucose)-ASN). The largest glycopeptides from Rous sarcoma virus were unbound to either Con A or lentil lectin and presumably contained tri- and/or tetra-antennary acidic-type structures ((NeuNAc +/- Gal-GlcNAc)3--4 -Man 3GlcNAc2 (+/- fucose)-ASN). In contrast, the majority of 'hybrid'-type oligosaccharides and essentially all of the neutral oligomannosyl core structures (Man5--9 GlcNAc1 and Man3 GlcNAc1) from the endoglycosidase-digested glycopeptides of both viruses were specifically bound to Con A-agarose, with the largest neutral oligosaccharides (Man7--9GlcNAc1) bound more tightly and less efficiently eluted by alpha-methyl mannoside.

Avian Sarcoma Viruses↗
PubMed1981

Transformation-dependent alterations in the oligosaccharides of Prague C Rous sarcoma virus glycoproteins.

The influence of cell transformation on the glycosylation of viral envelope glycoproteins was examined by high-resolution gel filtration and specific glycosidase digestions of 3H-sugar-labeled glycopeptides from nondefective and transformation-defective Prague C strains of Rous sarcoma virus replicated in fibroblasts from the same chicken embryo. The major difference in glycosylation attributable to the viral transformation of the host cells was an increase in this relative amount of larger acidic-type oligosaccharides containing additional "branch" sugars (NeuNAc-Gal-GlcNAc-) compared with the smaller acidic-type and neutral-type oligosaccharides. There was also a shift in size distribution of neutral-type oligosaccharides toward smaller oligomannosyl cores in the transforming versus nontransforming virus glycopeptides. These alterations were consistent with a transformation-dependent increase in the extent of intracellular processing of a common precursor structure for the asparagine-linked oligosaccharides of Rous sarcoma virus.

Animals↗
PubMed1981

Rous sarcoma virus glycoproteins contain hybrid-type oligosaccharides.

Examination of [3H]mannose-labeled glycopeptides from Prague C Rous sarcoma virus gp85 with gel filtration and sequential glycosidase digestions demonstrated the presence of hybrid-type asparaginyl-oligosaccharides. The major hybrid species had an oligomannosyl core (Man5GlcNAc2-ASN) characteristic of neutral structures, plus "branch" sugars (NeuNAc-Gal-GlcNAc-) characteristic of complex, acidic structures.

Asparagine↗
PubMed1980

CHO cells selected for phytohemagglutinin and con A resistance are defective in both early and late stages of protein glycosylation.

The lipid-linked and asparagine-linked oligosaccharides of two lectin-resistant and one parental Chinese hamster ovary (CHO) cell line have been compared by glycosidase digestion and gel filtration analysis of radiolabeled glycopeptides and oligosaccharides. The additional glycosylation defect in a double mutant cell line (CHO-PhaRConAR) selected from a phytohemagglutinin-resistant single mutant cell line (CHO-PhaR) for resistance to concanavalin A has been identified as a block in the synthesis of the lipid-linked oligosaccharide precursor, resulting in a structure with seven instead of the normal nine mannose units. Both the CHO-PhaRConR and CHO-PhaR cells were completely blocked in the synthesis of complex, acidic type oligosaccharides because of a previously demonstrated deficiency in a particular N--acetylglucosamine transferase activity. In addition, an altered collection of neutral type oligosaccharides (Man4-7GlcNAc2) accumulated in the glycoproteins of the double mutant.

Animals↗
PubMed1980

Altered synthesis and processing of oligosaccharides of vesicular stomatitis virus glycoprotein in different lectin-resistant Chinese hamster ovary cell lines.

To determine the particular intracellular steps in the glycosylation of the vesicular stomatitis virus (VSV) glycoprotein that were altered in several lectin-resistant CHO cell lines, VSV-infected parental and mutant cells were pulse-labeled for 30 and 120 min with [3H]mannose and [3H]glucosamine. Cell-associated viral glycopeptides were analyzed by gel filtration combined with specific glycosidase digestions and compared with the corresponding mature virion oligosaccharides. The intracellular glycosylation of the VSV glycoprotein in a mutant cell line resistant to phytohemagglutinin was identical to that in the normal cells except for a complete block in processing at a specific step in the final trimming of the oligomannosyl core from five to three mannoses. The results demonstrated that a double-mutant cell line selected from the phytohemagglutinin-resistant cells for resistance to concanavalin A had an additional defect in one of the earliest stages of glycosylation, resulting in smaller precursor oligosaccharides linked to protein.

Animals↗
PubMed1979

Sleep apnea, hypopnea and oxygen desaturation in normal subjects. A strong male predominance.

Thirty asymptomatic men and 19 asymptomatic women were monitored during one night's sleep to determine the incidence of breathing abnormalities and oxygen desaturation in normal subjects. Twenty men accounted for 264 episodes of nocturnal oxygen desaturation or abnormal breathing. Women never experienced oxygen desaturation, and only three had a total of nine episodes of apnea. These sex differences were highly significant (P less than 0.003). In men, increasing age and obesity correlated positively with the incidence of nocturnal oxygen desaturation and abnormal breathing. Four asymptomatic men weighing more than 90 kg dropped their saturation to very low levels (68 to 72 per cent). Abnormal breathing and oxygen desaturation during sleep in subjects with chronic obstructive lung disease of the syndrome of hypersomnolence with periodic breathing may represent the superimposition of smoking or obesity on a normal tendency to snoring and oxygen desaturation in men.

Adult↗
PubMed1979

Biosynthesis and maturation of cellular membrane glycoproteins.

The biosynthesis and the processing of asparagine-linked oligosaccharides of cellular membrane glycoproteins were examined in monolayer cultures of BHK21 cells and human diploid fibroblasts after pulse- and pulse-chase labeling with [2-3H]mannose. After pronase digestion, radiolabeled glycopeptides were characterized by high-resolution gel filtration, with or without additional digestion with various exoglycosidases and endoglycosidases. Pulse-labeled glycoproteins contained a relatively homogenous population of neutral oligosaccharides (major species: Man9GlcNAc2ASN). The vast majority of these asparagine-linked oligosaccharides was smaller than the major fraction of lipid-linked oligosaccharides from the cell and was apparently devoid of terminal glucose. After pulse-chase or long labeling periods, a significant fraction of the large oligomannosyl cores was processed by removal of mannose units and addition of branch sugars (NeuNAc-Gal-GlcNAc), resulting in complex acidic structures containing three and possibly five mannoses. In addition, some of the large oligomannosyl cores were processed by the removal of only several mannoses, resulting in a mixture of neutral structures with 5-9 mannoses. This oligomannosyl core heterogeneity in both neutral and acidic oligosaccharides linked to asparagine in cellular membrane glycoproteins was analogous to the heterogeneity reported for the oligosaccharides of avian RNA tumor virus glycoproteins (Hunt LA, Wright SE, Etchison JR, Summers DF: J Virol 29:336, 1979).

Acetylglucosaminidase↗
PubMed1979

Disordered breathing and oxygen desaturation during sleep in patients with chronic obstructive lung disease (COLD).

Seven patients with chronic obstructive lung disease (COLD) were monitored during their overnight sleep to determine the occurrence of disordered breathing and oxygen desaturation. Nasal and oral airflows were sensed by thermistor probes, chest wall movement by impedance pneumography and arterial oxygen saturation by ear oximetry. These variables were correlated with electroencephalographic and electrooculographic tracings. The subjects had a mean base line oxygen saturation of 89.2 per cent and slept an average of 218 minutes. Six of these seven subjects had one to 30 episodes of oxygen desaturation (decrease more than 4 per cent), 4 seconds to 30 minutes in duration, with declines in saturation as great as 36 per cent. In two subjects, saturation dropped to less than 50 per cent. Breathing was disordered in five of the seven subjects and included apnea and hypopnea. Subjects experienced from nine to 37 episodes of disordered breathing. Disordered breathing caused 42 per cent of the episodes of desaturation, all of which were less than 1 minute in duration. The mean maximum decline in saturation was 7.6 per cent. All episodes of desaturation lasting longer than 5 minutes occurred in rapid eye movement (REM) sleep and were not caused by disordered breathing. The mean maximal decrease in saturation was 22 per cent. This study reveals that disordered breathing is common in subjects with COLD and often causes desaturation but that it cannot explain all episodes of sleep desaturation.

Adult↗
PubMed1979

Oligosaccharide chains of avian RNA tumor virus glycoproteins contain heterogeneous oligomannosyl cores.

Chicken embryo fibroblasts (C/E phenotype) infected with subgroups B and C of the Prague strain of Rous sarcoma virus were radiolabeled with either [6-(3)H]-glucosamine or [2-(3)H]mannose, and virus was purified from the growth medium. The large envelope glycoprotein, gp85, was the only major radiolabeled component of purified virus. Pronase-digested glycopeptides from purified virus were analyzed by a combination of (i) gel filtration with columns of Sephadex G15/G50 and Bio-Gel P4 and (ii) enzymatic digestion of the oligosaccharide chains with specific exoglycosidases and endo-beta-N-acetylglucosaminidases. The rather broad molecular weight distribution (approximately 2,000 to 4,000) for glycopeptides in these studies and previous studies in other laboratories was shown to represent actual heterogeneity in the carbohydrate moieties: (i) the glycopeptides contained both mannose-rich, neutral chains and complex, acidic chains with terminal sialic acid; and (ii) both classes of asparagine-linked carbohydrate structures exhibited heterogeneity in the size of the oligomannosyl core (a mixture of approximately 5 to 9 mannose units for the neutral structures, and 3 or 5 mannose units for the acidic structures). With the [2-(3)H]mannose-labeled glycopeptides from Rous sarcoma virus, Prague strain subgroup C, most of the oligosaccharide chains were high-molecular-weight, acidic structures, with similar numbers of 3-mannose and 5-mannose core structures.

Acetylglucosaminidase↗
PubMed1979

Nocturnal pulmonary hypertension in patients with chronic obstructive pulmonary disease.

Oxygen desaturation occurs during sleep in some patients with COPD. To investigate the effects of these hypoxemic episodes on the pulmonary vasculature, we studied four patients with our routine polysomnographic techniques and simultaneously recorded pulmonary artery pressure. In all four subjects, nocturnal episodes of desaturation were accompanied by elevations in the pulmonary artery pressure. Low flow oxygen abolished the drops in arterial oxygen saturation (but not the breathing abnormalities) and no elevations in the PA pressure were observed. We postulate that in some COPD patients these initially transient events may lead to sustained pulmonary hypertension and cor pulmonale. Nocturnal oxygen therapy may be indicated in more patients than previously suspected and may prevent the development of cor pulmonale.

Adult↗
PubMed1978

Oligosaccharide chains are trimmed during synthesis of the envelope glycoprotein of vesicular stomatitis virus.

The biosynthesis and maturation of the oligosaccharide moieties of the envelope glycoprotein of vesicular stomatitis virus were investigated in virus-infected HeLa and BHK21 cells after pulse labeling with [2-3H]mannose. Two major forms of the virus glycoprotein were detected by polyacrylamide gel electrophoresis, which appear to correspond to the viral glycoprotein with either "precursor" or "mature" oligosaccharide chains. The precursor chains in both HeLa and BHK21 cells infected with vesicular stomatitis virus obtained after a 30-min pulse were large oligomannose structures containing approximately 7--9 mannose residues as estimated by gel filtration analysis. The size of the oligomannose structures initially transferred to the protein may have been even larger. Mature, virus-size oligosaccharide chains, which could be detected after a 20- to 30-min delay, contained only three mannose residues and, in addition, contained branch structures terminating in sialic acid. A precursor--product relationship of these two forms of oligosaccharide chains was demonstrated by pulse--chase labeling of virus-infected HeLa cells. These studies indicated that the large oligomannosyl core structures initially added to the glycoprotein were being "trimmed" by the removal of mannose residues prior to (and/or during) the addition of the branch chains terminating in sialic acid.

Cells, Cultured↗