Purification of rat adipose tissue lipoprotein lipase by affinity chromatography

Etienne, J; Breton, Michelyne; Vanhove, A.; Polonovski, J.

doi:10.1016/0005-2744(76)90042-5

Cited by 7 publications

(9 citation statements)

References 22 publications

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“…2) is generally similar to that described for the enzyme in other tissues. The only previous work on the rat adipose tissue enzyme is that of Greten & Walter (1978) and of Etienne et al (1976). Whereas Greten & Walter (1978) reported elution of the enzyme from heparin-Sepharose columns by 1.2M-NaCl as a single peak, in agreement with the results Fig.…”

Section: Discussionsupporting

confidence: 74%

“…reported here, Etienne et al (1976) described the elution of two peaks of activity with 1.16M-NaCl. Only the first of these, however, showed a requirement for serum in the assay medium and inhibition of the enzyme's activity in the assay by 1M-NaCl,…”

Section: Discussionmentioning

confidence: 86%

“…Such changes in the activity of the enzyme are believed to be hormonally determined and in recent years the control of the activity of the enzyme in rat adipose tissue by hormones has been a particular interest of this laboratory (Ashby et al, 1978b;Parkin et al, 1980). However, whereas the enzyme has been purified from human, porcine and avian adipose tissue, as well as from heart, milk and post-heparin plasma in a number of species (Nilsson-Ehle et al, 1980), there is little information on the molecular properties of the enzyme from rat adipose tissue (Greten & Walter, 1973;Etienne et al, 1976). In this paper we report the partial purification of the enzyme from this source, using the techniques of chromatography on heparin-Sepharose and concanavalin A-Sepharose that have been widely applied elsewhere, and provide evidence that the subunit of the enzyme, after separation by SDS/polyacrylamide-gel electrophoresis, has an apparent molecular weight of about 56000.…”

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confidence: 99%

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Purification and characterization of rat adipose tissue lipoprotein lipase

Parkin¹,

Speake²,

Robinson³

1982

Biochemical Journal

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Lipoprotein lipase (EC 3.1.1.34) extracted from adipose tissue of glucose-fed rats with 5 mM-sodium barbital, pH 7.5, containing 20% (v/v) glycerol and 0.1% (v/v) Triton X-100, was partially purified by affinity chromatography on heparin linked to Sepharose 4B. Sodium dodecyl sulphate/polyacrylamide-gel electrophoresis of the partially purified enzyme preparation revealed the presence of two major Coomassie-staining bands (mol.wts. 62 000 and 56 000) as well as a number of minor bands. Treatment of partially purified enzyme with [1,3-3H]di-isopropyl fluorophosphate resulted in the incorporation of radiolabel into the band of mol.wt. 56 000, but not into the band of mol.wt. 62 000. Both the amount of the 56 000-mol.wt. polypeptide and the incorporation of [1,3-3H]di-isopropyl fluorophosphate into this band were greatly reduced in the enzyme preparations isolated from adipose tissue of 48 h-starved rats. whereas the amount of the 62 000-mol.wt. polypeptide was unaffected by starvation. Purification of lipoprotein lipase from adipose tissue of glucose-fed rats was also carried out using affinity chromatography on Sepharose 4B linked to heparin with low affinity for antithrombin-III. This procedure resulted in the presence of a single band of mol.wt. 56 000 on sodium dodecyl sulphate/polyacrylamide-gel electrophoresis. These results suggest that the polypeptide of mol.wt. 56 000 corresponds to the subunit of lipoprotein lipase, whereas the 62 000-mol.wt. polypeptide probably represents antithrombin-III.

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Section: Discussionsupporting

confidence: 74%

Section: Discussionmentioning

confidence: 86%

mentioning

confidence: 99%

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Purification and characterization of rat adipose tissue lipoprotein lipase

Parkin¹,

Speake²,

Robinson³

1982

Biochemical Journal

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“…By heparin-Sepharose affinity chromatography, the soluble LPL of fat-pads could be dissociated into two active fractions. This has already been observed with acetone-dried powder of human or rat adipose tissues by several investigators (Etienne et al 1976;Garflnkel and Schotz l9'72;Davies, Oyer and Robinson 1974). It was reported by Vanhove, Wolf, Breton and Glangeaud (1978) that the first active fraction eluted from affinity chromatography was originated from the microsomal fractions, and the second one from the plasma membranes.…”

Section: Discussionmentioning

confidence: 59%

“…The column was equilibrated with 0.005 M barbital buffer, pH 8.6, containing 0.5 M NaCl and 20% glycerol at 4°C. Elution was carried out with a discontinuous NaCl gradient of increasing molarity (70 ml of 0.5 M; 70 ml of 0.7 M; 100 ml of 1.16 M) in 0.005 M sodium barbital buffer, pH 8.6, containing 20% glycerol (Etienne, Breton, Vanhove and Polonovski 1976). Fractions (5 ml) were collected and assayed for LPL activity.…”

Section: Separation Of Adipose Tissue Lipoprotein Lipasementioning

confidence: 99%

Effect of Pantethine on Post-Heparin Plasma Lipolytic Activities and Adipose Tissue Lipoprotein Lipase in Rats

Noma¹,

Kita²,

OKAMIYA³

1984

Horm Metab Res

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The lipid-lowering effect of pantethine, a new drug affecting lipid metabolism, had been evaluated in carbohydrate-induced hyperlipidemic rats. Administration of the drug raised post-heparin lipolytic activities, the change being due to an increase in lipoprotein lipase activity, whereas hepatic lipase activity remained virtually unchanged. Total lipoprotein lipase activity per g of adipose tissue increased in pantethine-treated rats compared with controls. Furthermore, the soluble lipoprotein lipase of fat-pads was fractionated by heparin-Sepharose affinity chromatography. The first active peak, originated from the microsomal fractions, significantly increased after the drug treatment, while the second one, originated from the plasma membranes, remained unchanged. The increase in the microsomal lipoprotein lipase activity may be due to an increase in intracellular synthesis of lipoprotein lipase enzyme proteins. The heterogeneity of lipoprotein lipase of rat adipose tissues was ensured using affinity chromatography on heparin-Sepharose.

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