scholarly journals Characterization of triacylglycerol hydrolase activities in isolated myocardial cells from rat heart

1985 ◽  
Vol 232 (1) ◽  
pp. 229-236 ◽  
Author(s):  
I Ramírez ◽  
A J Kryski ◽  
O Ben-Zeev ◽  
M C Schotz ◽  
D L Severson
Keyword(s):  
Ionic Strength ◽  
Subcellular Distribution ◽  
Rat Heart ◽  
High Ionic Strength ◽  
Hydrolase Activity ◽  
Affinity Column ◽  
Acid Hydrolase ◽  
Ph Optimum ◽  
Unbound Fraction ◽  
Triacylglycerol Hydrolase

Triacylglycerol (TG) hydrolase activities were characterized in myocytes isolated from rat hearts. Acid hydrolase activity with a pH optimum of 5 could be measured in myocyte homogenates, and the subcellular distribution suggested that this activity originated in lysosomes. Lipoprotein lipase (LPL) was also present in myocyte homogenates, as evidenced by TG hydrolase activity that was stimulated by serum and apolipoprotein CII, and inhibited by apolipoprotein CIII2, high ionic strength (NaCl and MgCl2, I = 1 M) and antibodies to LPL. Serum-independent neutral (pH 7.5) TG hydrolase activity was less sensitive to inhibition by 1 M-NaCl, by antibodies to LPL and by preincubation at 40 degrees C than was serum-stimulated hydrolase activity. Furthermore, there were modest but significant differences in the subcellular distribution of the serum-independent and serum-stimulated hydrolase activities. Hydrolase activities in myocyte homogenates could be solubilized by 7.2 mM-deoxycholate. Acid hydrolase activity was recovered in the unbound fraction after heparin-Sepharose chromatography, whereas LPL was bound to the affinity column and was eluted by 0.9-1.2 M-NaCl. Approximately one-third of the serum-independent TG hydrolase activity was not bound to the heparin-Sepharose affinity column. This unbound TG hydrolase activity had a pH optimum of 7 and was stimulated by 50 mM-MgCl2, but not by serum and was resistant to inhibition by high ionic strength (1 M-NaCl), to preincubation at 40 degrees C for 2 h, and by antibodies to LPL. It is concluded that, in addition to an acid lysosomal TG hydrolase and LPL, myocytes from rat heart contain a serum-independent TG hydrolase with unique characteristics.

scholarly journals Partial purification and properties of an AMP-specific soluble 5′-nucleotidase from pigeon heart

1990 ◽  
Vol 268 (1) ◽  
pp. 117-122 ◽  
Author(s):  
A C Skladanowski ◽  
A C Newby
Keyword(s):  
Ionic Strength ◽  
Molecular Mass ◽  
Formation Rate ◽  
Linear Increase ◽  
High Ionic Strength ◽  
Partial Purification ◽  
Ph Optimum ◽  
Purification And Properties ◽  
Wide Range ◽  
Alpha Beta

A soluble 5′-nucleotidase was purified 200-fold from pigeon heart. The enzyme (1) had an apparent molecular mass close to 150 kDa, (2) had a neutral pH optimum and hydrolysed a wide range of nucleoside 5′-monophosphates with a 15-fold preference for AMP over IMP, (3) at near-physiological concentrations of AMP was activated by ADP but not by ATP, (4) was inhibited by high Mg2+ concentration and high ionic strength, (5) was weakly inhibited by p-nitrophenol phosphate and Pi, and (6) was non-competitively inhibited more potently by 5′-deoxy-5′-isobutylthioinosine than by 5′-deoxy-5′-isobutylthioadenosine, but not by [alpha, beta-methylene]ADP. The data show that the enzyme is distinct from the ecto-5′-nucleotidase and from the previously purified IMP-specific 5′-nucleotidase. They also predict that the enzyme is activated during ATP catabolism and hence will generate a more-than-linear increase in the adenosine-formation rate in response to an increase in cytosolic AMP concentration.

scholarly journals Properties of mammalian nuclear-envelope nucleoside triphosphatase

1979 ◽  
Vol 181 (3) ◽  
pp. 647-658 ◽  
Author(s):  
P S Agutter ◽  
J B Cockrill ◽  
J E Lavine ◽  
B McCaldin ◽  
R B Sim
Keyword(s):  
Ionic Strength ◽  
Active Sites ◽  
Simian Virus 40 ◽  
Simian Virus ◽  
High Ionic Strength ◽  
3T3 Cells ◽  
Ph Optimum ◽  
Nucleoside Triphosphatase

The nucleoside triphosphatase activities of the nuclear envelopes from rat liver, pig liver and simian-virus-40-transformed mouse-embryo 3T3 cells were shown to exhibit similar parperties. All three preparations hydrolyse ATP, 2′-dATP, 3′-dATP, GTP, CTP and UTP in the presence of Mg2+, Ca2+, Mn2+ and Co2+ with a pH optimum of 8.0, are sensitive to inhibition by mercurials, arsenicals, quercetin, proflavin and adenosine 5′-[γ-thio]triphosphate and are partially inactivated by exposure to high ionic strength. The kinetic behaviour is similar for all substrates irrespective of the source of material. The typical Eadie-Hofstee plot, which is concave upwards at pH 8.0 when the ionic strength is 20mM, becomes linear when the pH is increased to 8.5 or the ionic strength to 160mM. The overall evidence, particularly the labelling of only one polypeptide by [gamma-32P]ATP, suggests that under the conditions of preparation and assay used only one class of nucleoside triphosphatase active sites is detectable in nuclear envelopes. The importance of these results for an understanding of the role of the enzyme in vivo is discussed.

Importance of Protease Inhibition in Studies on Purified Factor VIII (Antihaemophilic Factor)

1976 ◽  
Vol 35 (01) ◽  
pp. 186-190 ◽  
Author(s):  
Eugen A. Beck ◽  
Peter Bachmann ◽  
Peter Barbier ◽  
Miha Furlan
Keyword(s):  
Ionic Strength ◽  
Factor Viii ◽  
Gel Filtration ◽  
High Ionic Strength ◽  
Procoagulant Activity ◽  
Carrier Protein ◽  
Protease Inhibition ◽  
Functional Sites ◽  
Molecular Weight Peak ◽  
Von Willebrand

SummaryAccording to some authors factor VIII procoagulant activity may be dissociable from carrier protein (MW~ 2 × 106) by agarose gel filtration, e.g. at high ionic strength. We were able to reproduce this phenomenon. However, addition of protease inhibitor (Trasylol) prevented the appearance of low molecular weight peak of factor VIII procoagulant activity both at high ionic strength and elevated temperature (37°C). We conclude from our results that procoagulant activity and carrier protein (von Willebrand factor, factor VIII antigen) are closely associated functional sites of native factor VIII macro molecule. Consequently, proteolytic degradation should be avoided in functional and structural studies on factor VIII and especially in preparing factor VIII concentrate for therapeutic use.

THE RELATIVE DISTRIBUTION OF THYROXINE, TRIIODOTHYRONINE AND 3,3′,5′-(REVERSE)-TRIIODOTHYRONINE IN VARIOUS FRACTIONS OF THYROGLOBULIN

1978 ◽  
Vol 88 (2) ◽  
pp. 298-305 ◽  
Author(s):  
Peter Laurberg
Keyword(s):  
Ionic Strength ◽  
Guinea Pig ◽  
Sucrose Gradient ◽  
Deae Cellulose ◽  
High Ionic Strength ◽  
Relative Distribution ◽  
Thyroid Extract ◽  
Reverse Triiodothyronine ◽  
Thyroid Secretion ◽  
Stable Iodine

ABSTRACT Thyroglobulin fractions rich and poor in new thyroglobulin were separated by means of DEAE-cellulose chromatography of dog thyroid extracts and by zonal ultracentrifugation in a sucrose gradient of guinea pig thyroid extract incubated at low temperature. The distribution of thyroxine, triiodothyronine and 3,3′,5′-(reverse)-triiodothyronine in hydrolysates of the different fractions was estimated by radioimmunoassays. Following DEAE-cellulose chromatography there was a small but statistically significant increase in the T4/T3 ratio in thyroglobulin fractions eluted at high ionic strength - that is fractions relatively rich in stable iodine but poor in fresh thyroglobulin. There were no differences in the T4/rT3 ratios between the different fractions. The ratios between iodothyronines were almost identical in the various thyroglobulin fractions following zonal ultracentrifugation in a sucrose gradient of cold treated guinea pig thyroid extract. These findings lend no support to the possibility that a relatively high content of triiodothyronines in freshly synthesized thyroglobulin modulates the thyroid secretion towards a preferential secretion of triiodothyronine and 3,3′,5′-(reverse)-triiodothyronine at the expense of the secretion of thyroxine.

scholarly journals Kaolin based protective barrier in municipal landfills against adverse chemo-mechanical loadings

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Partha Das ◽  
Tadikonda Venkata Bharat
Keyword(s):  
Ionic Strength ◽  
High Ionic Strength ◽  
The Self ◽  
Design Parameters ◽  
Layered System ◽  
Geosynthetic Clay Liners ◽  
Clay Liners ◽  
Landfill Liner ◽  
First Time ◽  
Applied Stresses

AbstractIn this work, we assess the self-sealing and swelling ability of the compacted granular bentonite (GB) under an inorganic salt environment and induced overburden stresses from the landfill waste. The laboratory permeation tests with high ionic strength salt solutions reveal that the GB fails to seal and exhibits a significant mechanical collapse under different applied stresses. The applicability of GB in the form of geosynthetic clay liners as the bottom liner facilities in landfills that produce high ionic strength salt leachates, therefore, remains a serious concern. We propose an additional barrier system based on kaolin, for the first time, to address this problem. The proposed kaolin-GB layered system performs satisfactorily in terms of its sealing and swelling ability even in adverse saline conditions and low overburden stresses. The kaolin improves the osmotic efficiency of the self and also helps the underlying GB layer to seal the inter-granular voids. The estimated design parameters by through-diffusion test suggest that the kaolin-GB layered system effectively attenuates the permeant flux and suitable as a landfill liner.

scholarly journals High ionic strength dissociation assay (HISDA) for high drug tolerant immunogenicity testing

Bioanalysis ◽  
2020 ◽  
Author(s):  
Gregor Jordan ◽  
Alexander Pöhler ◽  
Florence Guilhot ◽  
Meike Zaspel ◽  
Roland F Staack
Keyword(s):  
Ionic Strength ◽  
Acid Treatment ◽  
Structural Integrity ◽  
Drug Tolerance ◽  
High Ionic Strength ◽  
Acid Dissociation ◽  
High Drug ◽  
Overnight Incubation ◽  
Antidrug Antibody ◽  
High Doses

Aim: Antidrug antibody (ADA) assessment may be challenged in studies that involve the administration of high doses of biotherapeutics and/or with long half-lives. In such cases, ADA assays with optimized drug tolerance are desired. Material & Methods: We evaluated the use of MgCl2 to develop high ionic strength dissociation assays in two investigational examples (bridging enzyme-linked immunosorbent ADA assays) to attain high drug tolerance while maintaining best possible structural integrity of ADAs. Results: Both ADA-bridging assays treated with MgCl2 showed improved drug tolerance and higher signal-to-blank values compared with overnight incubation or acid treatment. Conclusion: The use of MgCl2 treatment in ADA-bridging assays provides a sensitive, drug tolerant and easy-to-use alternative in cases where acid dissociation is not possible or unwanted.

scholarly journals High salt compatible oxyanion receptors by dual ion imprinting

2020 ◽  
Vol 11 (16) ◽  
pp. 4246-4250 ◽  
Author(s):  
Sudhirkumar Shinde ◽  
Mona Mansour ◽  
Anil Incel ◽  
Liliia Mavliutova ◽  
Celina Wierzbicka ◽  
...  
Keyword(s):  
Ionic Strength ◽  
Ion Pair ◽  
High Ionic Strength ◽  
High Salt ◽  
Ion Uptake ◽  
Ion Imprinting

Imprinting of an ion-pair in presence of mutually compatible anion and cation host monomers leads to polymers showing enhanced ion uptake in competitive high ionic strength buffers.

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