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A Lagrou

Publications and source records attributed to A Lagrou.

At least 37 records · Page 2Linked to original sources
PubMed1985

Fluidity characteristics of bovine thyroid plasma membranes.

Highly purified plasma membranes of bovine thyroid were obtained by differential pelleting followed by discontinuous gradient centrifugation in a swing-out rotor. Subfractions of plasma membranes were prepared by affinity chromatography on Con A-Sepharose. The final membrane fractions were enriched 25-30-fold over homogenate in 5'-nucleotidase and alkaline phosphatase and displayed a protein to phospholipid ratio of 1.67 and a cholesterol to phospholipid molar ratio of 0.55. The phospholipid composition did not deviate appreciably from that of whole tissue except for the higher sphingomyelin level (22.5 vs. 14.0%). The predominant fatty acids were palmitic (16:0), oleic (18:1), stearic (18:0) and linoleic (18:2) acid. The physical state of the membrane was studied by (i) calculation of the lipid structural order parameter SDPH from steady-state fluorescence anisotropy determinations of the hydrophobic probe 1,6-diphenyl-1,3,5-hexatriene (DPH); (ii) estimation of the lateral diffusion coefficient of pyrene following excimer formation. These parameters were determined in native thyroid plasma membranes and in reconstituted vesicles, obtained by detergent dialysis from octylglucoside solubilized membrane components. The presence of membrane protein or neutral lipids induced more restraint on the movements of the fluorophores. The lipid order parameter, SDPH was mainly determined by the neutral lipids. Subfractions of plasma membrane enriched in luminal membranes have a slightly lower fluidity (higher SDPH and lower Ddiff values) than subfractions enriched in basolateral membranes. This difference appears to be due to both differences in lipid as well as protein composition. Under physiological conditions, no significant alterations in probe dynamics could be observed upon addition of thyrotropin or cholera toxin, even at micromolar concentrations.

Animals↗
PubMed1984

Identification and characterization of dolichyl dolichoate, a novel isoprenoic derivative in bovine thyroid.

Inspection of the TLC pattern of the neutral lipid fraction of bovine thyroid reveals, in addition to cholesteryl esters and dolichyl fatty acid esters, the presence of a not yet identified compound in the most apolar lipid region. This unknown compound was purified on a preparative scale by silicic acid column chromatography, Lipidex-5000 gel-filtration chromatography and preparative thin-layer chromatography. By chemical (hydrolysis and reduction) and spectroscopic (ultraviolet, infrared, NMR, mass spectroscopy) methods this lipid was identified as dolichyl dolichoate. The homologue pattern was analyzed, by HPLC and FDMS, respectively.

Animals↗
PubMed1984

Spectrophotometric determination of dolichol and dolichyl derivatives using the Chugaev color reaction.

A simple colorimetric method for the assay of microamounts of dolichol is described. It is based essentially on the Chugaev color reaction. The procedure allows the rapid (1 h after purification of the sample, less than half a day for the whole procedure) and reliable (% SE less than or equal to 5%) determination of dolichol and dolichyl derivatives in the microgram range. Color production is linear within the concentration range 2 to 20 micrograms dolichol. With biological samples, prior to colorimetric assay, dolichol and dolichyl derivatives are isolated by solvent extraction and TLC. Recoveries are monitored by isotope dilution analysis. The method has been used for the analysis of dolichol and dolichyl ester levels in bovine thyroid tissue. The procedure can also be applied for the analysis of other isoprenoids.

Animals↗
PubMed1984

A study on the RNA polymerase activities in bovine thyroid nuclei isolated in isotonic sucrose, hypertonic sucrose or in anhydrous glycerol media. Influence of the isolation procedure on the electrophoretic pattern of acid soluble nuclear proteins.

After differential pelleting of bovine thyroid bound RNA polymerase II was the more enriched enzyme activity in the nuclear fraction, and coincided best with the DNA profile. The RNA polymerase I + III activity was compared in nuclear fractions isolated either in 0.25 M sucrose (wet tissue) or in anhydrous glycerol (lyophilized tissue) or in 2.4 M sucrose (lyophilized tissue). Although the nuclei were more resistant to the isolation procedure in glycerol, more proteins were extracted by that procedure than during the isolation in 2.4 M sucrose. With the 2.4 M sucrose method a twofold enrichment of RNA polymerase I + III activity in respect to DNA occurred in the nuclei pointing to an exclusive localization of these activities within the nucleus. Using the same isolation procedure the different classes of histones were better resolved upon polyacrylamide gel electrophoresis.

Animals↗
PubMed1984

Characterization, purification, and subcellular localization of bovine thyroid sialidases.

Sialidase activities have been studied in bovine thyroid using sialoglycolipids, sialoglycoproteins, sialo-oligosaccharides and fluorogenic 4-methylumbelliferyl-alpha-D-N-acetylneuraminate as substrates. No sialidase activity could be detected towards native glycoprotein substrates. From enzyme kinetics, effector data and more convincingly from subcellular studies it became clear that in bovine thyroid at least two sialidase activities were present, a sialyllactitol sialidase confined to the lysosomal membrane and a glycolipid sialidase residing in the plasma membrane and displaying the features of a true ectoenzyme. The lipid requirement for full enzyme activity supported the membrane bound character of both sialidase activities. A soluble sialidase activity could not be demonstrated. After solubilization by CHAPS treatment, partial purification of the sialyllactitol sialidase could be achieved by affinity chromatography (Sepharose diamino dipropylamino-N-acetylneuraminic acid). The purified enzyme was extremely labile. Titration of the sialidase preparation with amino acid modifying agents revealed that sulfhydryl- and tryptophanyl groups were essential for the sialidase action.

Animals↗
PubMed1982

DNA-dependent RNA polymerases from bovine thyroid: catalytic properties and template specificities.

1. DNA-dependent RNA polymerases I and II have been purified starting from bovine thyroid nuclei yielding a purification factor of 230 for the RNA polymerase I and a purification factor 3212 for RNA polymerase II. RNA polymerase II was further characterized by gel electrophoresis and amino-acid analysis. 2. Kinetics and optimal assay conditions for both RNA polymerases were studied. 3. The template efficiency of a number of DNA preparations was investigated. 4. Rifamycin AF 013 and heparin act as initiation inhibitors. 5. Polyamines were shown to enhance the rate of chain elongation.

Animals↗
PubMed1980

The subcellular biochemistry of thyroid.

In this review the subcellular localization of enzymes and constituents in thyroid is discussed. Conditions and results of differential pelleting and gradient centrifugation studies are described with special attention to the validity of the markets used (Table VI). Special approaches to the isolation and characterization of thyroid organelles and membranes are extensively reviewed (Table VII). Subcellular fractionation of thyroid tissue has been shown to be an arduous task. Classic approaches for differential pelleting and gradient centrifugation, which have been proved successful for rat liver, are not always equally satisfactory for thyroid. The major problem is the toughness of the tissue requiring rather traumatizing homogenizing procedures. Nevertheless, the fractionation procedures did allow the subcellular localization of some enzymes and constituents to be established with a high degree of certainty. Furthermore, enriched subcellular fractions have been isolated which have been useful for biochemical studies concerning the specific function of this tissue.

Animals↗
PubMed1979

Isolation and identification of polyprenols from bovine thyroid gland.

The presence of polyprenols in bovine thyroid was demonstrated. After preparative isolation, the structure was elucidated by chemical and spectroscopic techniques. The main polyprenol homologue has a molecular weight of 1380 corresponding to the presence of 20 isoprene units. From NMR studies it appears that 18 units have the cis configuration and that the 2 others are trans isoprene units. The dolichol content amounts to 0.2 mg/g wet weight. About 5% was found in the esterified form.

Animals↗
PubMed1979

Two cases of mucopolysaccharidosis type III (Sanfilippo). An anatomopathological study.

Anatomopathological studies of one case of Sanfilippo disease types A and C, respectively, are presented. The storage phenomenon is very severe in the central as well as in the peripheral nervous system of both patients. Light microscopy does not show significant differences between the two cases. Electron microscopy of the nervous system reveals in both cases the presence of variable amounts of zebra bodies and of membrano-granulo-vacuolar inclusions. The presence of larger amounts of zebra bodies in the type A case and of larger quantities of membrano-granulo-vacuolar inclusions in the type C case constitute probably a nondistinctive feature between the two types. Light and electron microscopic studies of visceral organs do not disclose significant differences either. It is concluded that no major morphological differences between Sanfilippo disease types A and C can be observed.

Adolescent↗
PubMed1979

Two cases of mucopolysaccharidosis type III (Sanfilippo). A biochemical study.

The mucopolysaccharide and lipid composition of human nervous tissue and viscera from one case of Sanfilippo disease type A and one case of Sanfilippo disease type C, were investigated. In the brain a moderate increase of acid glycosaminoglycans occurred. This phenomenon was much more pronounced in the viscera, especially in the liver. In all tissues this increase was mainly due to an accumulation of heparan sulphate. Changes in lipid composition were noted, but can be regarded as secondary effects. The biochemical results reported also suggest some general conclusions. (a) AGAG and lipid analyses do not permit differentiation between the subtypes of Sanfilippo disease. (b) The differences in lipid composition can probably be considered as consequences of variation in secondary effects. (c) The severe demyelination in brain correlates well with the biochemical lipid analysis. However, in other instances it remains difficult to bridge the gap still existing between some morphological and biochemical data.

Adolescent↗
PubMed1979

RNA synthesis in isolated bovine thyroid nuclei and nucleoli. alpha-Amanitin effect, a hint to the existence of a specific regulatory system.

DNA dependent RNA polymerase activities in isolated bovine thyroid nuclei and nucleoli have been studied. They retain their RNA synthetic activity for an extended period of time. This RNA synthetic activity is sensitive to actinomycin D and requires the presence of all four ribonucleoside triphosphates. The optimal conditions have been determined. Polyacrylamide gel electrophoresis reveals that the RNA synthesized has a size distribution ranging from 34S to 4S. The production of 18S-8S RNA is very sensitive to low concentrations of alpha-amanitin. However, in isolated bovine thyroid nuclei (not in nucleoli) this drug displays an effect on all RNA classes produced. The alpha-amanitin induced drastic decrease of [3H]-UMP incorporation in RNA of all sizes synthesized by isolated bovine thyroid nuclei is discussed.

Amanitins↗
PubMed1979

Lipolytic enzymes in bovine thyroid tissue. III. Lysophospholipase activity.

Lysophospholipids are formed during phospholipid breakdown as a result of the action of phospholipases A. At certain concentrations these lysoderivatives destabilise biological membranes. Therefore, their concentration is of critical importance for membrane integrity. Prevention of lysophosphoglycerides accumulation may be the important role for lysophospholipases and is probably the explanation for their widespread occurrence in nature. Lysophospholipase activities were found in molds (Fairbairn, 1948), rice bran (Contardi & Ercoli, 1933), several microorganisms (Brockerhoff & Jensen, 1974), snake and bee venoms (Doery & Pearson, 1964; Mohamed et al., 1969; Shiloah et al., 1973), insects (Khan & Hodgson, 1967; Rao & Subrahmanyam, 1969), fish muscle (Yurkovski & Brockerhoff, 1965; Cohen et al., 1967) and in various animal tissues (Marples & Thompson, 1960). In mammalian tissue the enzyme was first described in beef pancreas (Shapiro, 1953). Relatively high levels were detected in intestine, lung, spleen, liver and pancreas, while lower levels were present in muscle, kidney, testes, brain and blood (Marples & Thompson, 1960). The presence of lysophospholipase activity in both supernatant and sediment of bovine thyroid was reported previously in relation to possible interference of this enzyme with the phospholipase A activity assay (De Wolf et al., 1976). The subcellular localization of bovine thyroid lysophospholipase and some properties of the membrane bound enzyme activity are discussed in this paper.

Animals↗
PubMed1979

Occurrence and subcellular localization of glucose 6-phosphatase in bovine thyroid.

In bovine thyroid tissue the glucose 6-phosphatase activity is not entirely due to the presence of an unspecific acid phenylphosphatase nor to beta-glycerophosphatase. This glucose 6-phosphatase is very probably localized within endoplasmic reticulum membranes. It is not a good marker for distribution patterns obtained after differential pelleting. However it can be used as a marker for endoplasmic reticulum membranes after centrifugation in a zonal rotor.

Animals↗