scholarly journals Biochemical studies on the isolation and characterization of human spleen haemosiderin

1984 ◽  
Vol 223 (1) ◽  
pp. 31-38 ◽  
Author(s):  
M P Weir ◽  
J F Gibson ◽  
T J Peters
Keyword(s):  
Minor Component ◽  
Iron Oxyhydroxide ◽  
Human Spleen ◽  
Major Band ◽  
Isolation And Characterization ◽  
Biochemical Studies ◽  
Characteristic Group ◽  
A Minor ◽  
Proteolytic Product

Haemosiderin was isolated from thalassaemic human spleens by centrifugation through concentrated KI solutions. A method for solubilizing haemosiderin was developed which leaves the iron oxyhydroxide cores and constituent polypeptides intact, facilitating further purification and analysis. Purified haemosiderin contained no detectable haem, trace amounts of carbohydrate, and iron and phosphorus in a molar ration of 6:1; much of the phosphate may be present as core-adsorbed. Several lipids were present, but it is not certain whether these are contaminants or components of the haemosiderin granules. In all preparations examined, a characteristic group of six to seven peptides of apparent Mr 12 900-17 800 were found, with a major band at Mr 14 500 and, in addition, a minor component of Mr 42 000; these peptides co-chromatographed with the cores. Negatively stained electron micrographs suggest that these peptides form an incomplete shell about the cores, consistent with the view that haemosiderin is a proteolytic product of ferritin.

Analysis of plant genomes. IV. Isolation and characterization of satellite DNA components from two dicotyledons cucumber (Cucumis sativus) and radish (Raphanus sativus)

1978 ◽  
Vol 56 (8) ◽  
pp. 808-815 ◽  
Author(s):  
P. K. Ranjekar ◽  
D. Pallotta ◽  
J. G. Lafontaine
Keyword(s):  
Satellite Dna ◽  
Raphanus Sativus ◽  
Nuclear Dna ◽  
Minor Component ◽  
Isolation And Characterization ◽  
Dna Contents ◽  
A Minor ◽  
Dna Components ◽  
Denaturation Profile

Satellite DNA fractions from cucumber and radish, two plants having low DNA contents and relatively small chromosomes, were isolated and characterized. Reassociation studies of satellite and total nuclear DNA showed that the satellite fractions in these two plants contain most of the rapidly reassociating DNA. Cucumber satellite I was found to contain one major component (70% of the total satellite) having a density of 1.706 g/cm3 and a Tm of 90.5 °C and a minor component with a density of 1.712 g/cm3 and a Tm of 93.5 °C. The complexity of the major component was estimated to be 3.8 × 105 daltons while that of the minor one was 12.9 × 107 daltons. Although cucumber satellite II banded as a single peak at adensity of 1.700 g/cm3 in neutral CsCl gradients, it was observed to have a rather broad denaturation profile with a Tm of 86.5 °C. Its Cot curve was also broader than that of satellite I and one of its components (40% of the total) had a complexity of 5.8 × 105 daltons. Two satellite fractions were also observed in the case of radish DNA but only satellite I was isolated in a pure form and characterized. This radish satellite formed a sharp, symmetrical peak at a density of 1.698 g/cm3 in neutral CsCl gradients and underwent denaturation in a narrow temperature range of 6 to 7 °C. An analysis of the optical reassociation kinetics showed that this satellite contained a major and a minor component. The major component, which comprised 80% of the satellite, had a complexity of 12.9 × 105 daltons. Hybridization experiments revealed that the ribosomal DNA was present in satellite II.

Functional Characterization of GABAA Receptors in Neonatal Hypothalamic Brain Slice

2002 ◽  
Vol 88 (4) ◽  
pp. 1655-1663 ◽  
Author(s):  
Ren-Qi Huang ◽  
Glenn H. Dillon
Keyword(s):  
Functional Characterization ◽  
Minor Component ◽  
Dehydroepiandrosterone Sulfate ◽  
Receptor Subtypes ◽  
Hypothalamic Neurons ◽  
Inhibitory Neurotransmitter ◽  
Old Rats ◽  
A Minor ◽  
Whole Cell Recordings

The hypothalamus influences a number of autonomic functions. The activity of hypothalamic neurons is modulated in part by release of the inhibitory neurotransmitter GABA onto these neurons. GABAA receptors are formed from a number of distinct subunits, designated α, β, γ, δ, ε, and θ, many of which have multiple isoforms. Little data exist, however, on the functional characteristics of the GABAA receptors present on hypothalamic neurons. To gain insight into which GABAA receptor subunits are functionally expressed in the hypothalamus, we used an array of pharmacologic assessments. Whole cell recordings were made from thin hypothalamic slices obtained from 1- to 14-day-old rats. GABAA receptor-mediated currents were detected in all neurons tested and had an average EC50 of 20 ± 1.6 μM. Hypothalamic GABAA receptors were modulated by diazepam (EC50 = 0.060 μM), zolpidem (EC50 = 0.19 μM), loreclezole (EC50 = 4.4 μM), methyl-6,7-dimethoxy-4-ethyl-β-carboline (EC50= 7.7 μM), and 5α-pregnan-3α-hydroxy-20-one (3α-OH-DHP). Conversely, these receptors were inhibited by Zn2+ (IC50 = 70.5 μM), dehydroepiandrosterone sulfate (IC50 = 16.7 μM), and picrotoxin (IC50 = 2.6 μM). The α4/6-selective antagonist furosemide (10–1,000 μM) was ineffective in all hypothalamic neurons tested. The results of our pharmacological analysis suggest that hypothalamic neurons express functional GABAA receptor subtypes that incorporate α1 and/or α2 subunits, β2 and/or β3 subunits, and the γ2 subunit. Our results suggest receptors expressing α3–α6, β1, γ1, and δ, if present, represent a minor component of functional hypothalamic GABAA receptors.

Genetic and biochemical studies of asparagine-linked oligosaccharide assembly

1982 ◽  
Vol 300 (1099) ◽  
pp. 207-223 ◽  
Keyword(s):  
Protein Synthesis ◽  
Temperature Sensitive ◽  
Temperature Sensitive Mutants ◽  
Isolation And Characterization ◽  
Oligosaccharide Processing ◽  
Biochemical Studies ◽  
The Common ◽  
Specific Lipid ◽  
Temperature Sensitive Phenotype

The formation of N -glycosidic linkages of eukaryotic glycoproteins involves the assembly of a specific lipid-linked precursor oligosaccharide in the endoplasmic reticulum. This oligosaccharide is transferred from the lipid carrier to appropriate asparagine residues during protein synthesis. The protein-linked oligosaccharide then undergoes processing reactions that include both removal and addition of carbohydrate residues. In this paper we report recent studies from our laboratory on the synthesis of asparagine-linked oligosaccharides. In the first part we describe the isolation and characterization of temperature-sensitive mutants of yeast blocked at specific stages in the assembly of the lipid-linked oligosaccharide. In addition, we are using these mutants to clone the genes for the enzymes in this pathway by complementation of the temperature-sensitive phenotype. The second part deals with the topography of asparagine-linked oligosaccharide assembly. Our studies on the transmembrane movement of sugar residues during the assembly of secreted glycoproteins from cytoplasmic precursors are presented. Finally, experiments on the control of protein-linked oligosaccharide processing are described. Recent data are presented on the problem of how specific oligosaccharides are assembled from the common precursors at individual sites on glycoproteins.

Biochemical Studies on Sphingolipid of Artemia franciscana (I) Isolation and Characterization of Sphingomyelin

Lipids ◽  
2010 ◽  
Vol 45 (7) ◽  
pp. 635-643 ◽  
Author(s):  
Hisao Kojima ◽  
Takashi Inoue ◽  
Mutsumi Sugita ◽  
Saki Itonori ◽  
Masahiro Ito
Keyword(s):  
Artemia Franciscana ◽  
Isolation And Characterization ◽  
Biochemical Studies

scholarly journals Isolation and characterization of a minor fimbria from Porphyromonas gingivalis.

1996 ◽  
Vol 64 (11) ◽  
pp. 4788-4794 ◽  
Author(s):  
N Hamada ◽  
H T Sojar ◽  
M I Cho ◽  
R J Genco
Keyword(s):  
Porphyromonas Gingivalis ◽  
Isolation And Characterization ◽  
A Minor

scholarly journals Characterization of Heat-Inducible Expression and Cloning of HtpG (Hsp90 Homologue) of Porphyromonas gingivalis

2000 ◽  
Vol 68 (4) ◽  
pp. 1980-1987 ◽  
Author(s):  
Dennis E. Lopatin ◽  
Allison Combs ◽  
Domenica G. Sweier ◽  
J. Christopher Fenno ◽  
Sangeeta Dhamija
Keyword(s):  
Porphyromonas Gingivalis ◽  
Cellular Localization ◽  
Single Gene ◽  
Serum Antibody ◽  
Stress Protein ◽  
Major Band ◽  
Disease Associations ◽  
A Minor

ABSTRACT Porphyromonas gingivalis is implicated in the etiology of periodontal disease. Associations between microbial virulence and stress protein expression have been identified in other infections. For example, Hsp90 homologues in several microbial species have been shown to contribute to virulence. We previously reported that P. gingivalis possessed an Hsp90 homologue (HtpG) which cross-reacts with human Hsp90. In addition, we found that elevated levels of serum antibody to Hsp90 stress protein in individuals colonized with this microorganism were associated with periodontal health. However, the role of HtpG in P. gingivalis has not been explored. Therefore, we cloned the htpG gene and investigated the characteristics of HtpG localization and expression in P. gingivalis. htpG exists as a single gene of 2,052 bp from which a single message encoding a mature protein of approximately 68 kDa is transcribed. Western blot analysis revealed that the 68-kDa polypeptide was stress inducible and that a major band at 44 kDa and a minor band at 40 kDa were present at constitutive levels. Cellular localization studies revealed that the 44- and 40-kDa species were associated with membrane and vesicle fractions, while the 68-kDa polypeptide was localized to the cytosolic fractions.

scholarly journals Purification and Characterization of Thin Pili of IncI1 Plasmids ColIb-P9 and R64: Formation of PilV-Specific Cell Aggregates by Type IV Pili

1998 ◽  
Vol 180 (11) ◽  
pp. 2842-2848 ◽  
Author(s):  
Tetsu Yoshida ◽  
Nobuhisa Furuya ◽  
Masayuki Ishikura ◽  
Toshiaki Isobe ◽  
Kazu Haino-Fukushima ◽  
...  
Keyword(s):  
Gradient Centrifugation ◽  
Minor Component ◽  
Electrophoretic Analysis ◽  
Type Iv Pili ◽  
Specific Cell ◽  
Cell Aggregates ◽  
Type Iv ◽  
Cscl Density Gradient ◽  
A Minor

ABSTRACT Thin pili of the closely related IncI1 plasmids ColIb-P9 and R64 are required only for liquid mating and belong to the type IV family of pili. They were sedimented by ultracentrifugation from culture medium in which Escherichia coli cells harboring ColIb-P9- or R64-derived plasmids had been grown, and then the pili were purified by CsCl density gradient centrifugation. In negatively stained thin pilus samples, long rods with a diameter of 6 nm, characteristic of type IV pili, were observed under an electron microscope. Gel electrophoretic analysis of purified ColIb-P9 thin pili indicated that thin pili consist of two kinds of proteins, pilin and the PilV protein. Pilin was demonstrated to be the product of the pilS gene. Pilin was first synthesized as a 22-kDa prepilin from the pilS gene and subsequently processed to a 19-kDa protein by the function of thepilU product. The N-terminal amino group of the processed protein was shown to be modified. The C-terminal segments of thepilV products vary among six or seven different types, as a result of shufflon DNA rearrangements of the pilV gene. These PilV proteins were revealed to comprise a minor component of thin pili. Formation of PilV-specific cell aggregates by ColIb-P9 and R64 thin pili was demonstrated and may play an important role in liquid mating.

Further purification and characterization of newly synthesized anionic glycoconjugates secreted by cultured UMR-106 cells: evidence that the major anionic glycoconjugate secreted by these cells is similar to bone sialoprotein II

1991 ◽  
Vol 69 (8) ◽  
pp. 523-530 ◽  
Author(s):  
Ravi K. Chopra ◽  
Tassos P. Anastassiades ◽  
David Lohnes ◽  
Glenville Jones
Keyword(s):  
Amino Acid ◽  
Cetylpyridinium Chloride ◽  
Polyacrylamide Gel Electrophoresis ◽  
Minor Component ◽  
Keratan Sulfate ◽  
Bone Sialoprotein ◽  
Terminal Amino ◽  
A Minor ◽  
Umr 106

Following incubation of UMR-106 cells for 48 h in the presence of [3H]glucosamine and [35S]sulfate, the newly synthesized anionic glycoconjugates were isolated from the culture medium by cetylpyridinium chloride/ethanol precipitation and further separated by DEAE-Sephacel chromatography into two radiolabelled fractions, a major component, UM I, and a minor component, UM II. UM I appeared to be homogeneous as shown by Sepharose CL-4B chromatography under dissociative conditions, and SDS-polyacrylamide gel electrophoresis. It showed a molecular mass of approximately 93 kDa on 4–15% gels. UM I was partially degraded by brief treatment with trypsin, releasing a small, terminal peptide that contained 47.6% of 35S but no 3H. Treatment of UM I with neuraminidase and 0.1 N H2SO4 (1 h at 80 °C), respectively, released 27% 3H and 38.4% 3H plus 41% 35S, suggesting the presence of a significant number of sialic acid residues, as shown by Sephadex G-50 chromatography of the digests. Amino acid analysis showed that the UM I glycoconjugate was rich in acidic amino acids (12.6% aspartic acid and 21.2% glutamic acid residues) and its N-terminal sequence was Phe-Ser-Met-Lys-Asn-Phe-, which is identical to the published N-terminal amino acid sequence of rat bone sialoprotein II. Keratanase treatment of UM I released 26% of the incorporated radioactivity, suggesting the presence of keratan sulfate chains. UM II contained a chondroitinase ABC-sensitive proteoglycan.Key words: UMR-106 cells, anionic glycoconjugates, bone sialoprotein II.

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