Phospholipid metabolism in the molluscs. II. Activities of choline kinase, ethanolamine kinase, and CTP:phosphorylethanolamine cytidyltransferase in the mollusc Helix lactea

1970 ◽  
Vol 48 (5) ◽  
pp. 580-584 ◽  
Author(s):  
Chi-Rong Liang ◽  
M. Segura ◽  
K. P. Strickland
Keyword(s):  
Digestive Gland ◽  
High Speed ◽  
Enzyme Preparation ◽  
Phospholipid Metabolism ◽  
Maximum Activity ◽  
Supernatant Fraction ◽  
Choline Kinase ◽  
Lipid Phosphorus ◽  
Ethanolamine Kinase ◽  
High Speed Supernatant

The distribution of phospholipids in the mollusc Helix lactea was investigated. The results showed that ethanolamine and choline phosphoglycerides are the major phospholipids in this species, amounting to 49% and 26% of the total lipid phosphorus. The amount of ceramide 2-aminoethylphosphonate was 9% of the total phospholipids and this compound was the only phosphonolipid detected in this species. Ethanolamine and choline kinase activities were shown to be present in the high-speed supernatant fraction of snail digestive gland. The maximum activity of choline kinase was found to be higher than that of ethanolamine kinase in the same enzyme preparation. The enzyme CTP:phosphorylethanolamine cytidyltransferase was also demonstrated in the high-speed supernatant fraction of snail digestive gland. The enzymic reaction product was identified as CDP-ethanolamine by paper chromatography and radioautography. The enzymes phosphorylcholine cytidyltransferase and aminoethylphosphonate cytidyltransferase were not detected under the conditions employed in the phosphorylethanolamine cytidyltransferase assays.

scholarly journals Detection of CMP-N-acetylneuraminic acid hydroxylase activity in fractionated mouse liver

1989 ◽  
Vol 263 (2) ◽  
pp. 355-363 ◽  
Author(s):  
L Shaw ◽  
R Schauer
Keyword(s):  
Submandibular Gland ◽  
Mouse Liver ◽  
High Speed ◽  
Metal Salts ◽  
Supernatant Fraction ◽  
Inhibitory Influence ◽  
Hydroxylase Activity ◽  
Acetylneuraminic Acid ◽  
Liver Homogenates ◽  
High Speed Supernatant

The finding that N-glycoloylneuraminic acid (Neu5Gc) in pig submandibular gland is synthesized by hydroxylation of the sugar nucleotide CMP-Neu5Ac [Shaw & Schauer (1988) Biol. Chem. Hoppe-Seyler 369, 477-486] prompted us to investigate further the biosynthesis of this sialic acid in mouse liver. Free [14C]Neu5Ac, CMP-[14C]Neu5Ac and [14C]Neu5Ac glycosidically bound by Gal alpha 2-3- and Gal alpha 2-6-GlcNAc beta 1-4 linkages to fetuin were employed as potential substrates in experiments with fractionated mouse liver homogenates. The only substrate to be hydroxylated was the CMP-Neu5Ac glycoside. The product of the reaction was identified by chemical and enzymic methods as CMP-Neu5Gc. All of the CMP-Neu5Ac hydroxylase activity was detected in the high-speed supernatant fraction. The hydroxylase required a reduced nicotinamide nucleotide [NAD(P)H] coenzyme and molecular oxygen for activity. Furthermore, the activity of this enzyme was enhanced by exogenously added Fe2+ or Fe3+ ions, all other metal salts tested having a negligible or inhibitory influence. This hydroxylase is therefore tentatively classified as a monooxygenase. The cofactor requirement and CMP-Neu5Ac substrate specificity are identical to those of the enzyme in high-speed supernatants of pig submandibular gland, suggesting that this is a common route of Neu5Gc biosynthesis. The relevance of these results to the regulation of Neu5Gc expression in sialoglycoconjugates is discussed.

CHOLINE KINASE OF RAPESEED (BRASSIGA CAMPESTRIS L.)

1957 ◽  
Vol 35 (1) ◽  
pp. 853-863
Author(s):  
T. Ramasarma ◽  
L. R. Wetter
Keyword(s):  
Calcium Phosphate ◽  
Enzyme Preparation ◽  
Brassica Campestris ◽  
Maximum Activity ◽  
Choline Kinase ◽  
Aqueous Extracts ◽  
Neutral Ph ◽  
Sulphydryl Groups

Choline kinase from aqueous extracts of Polish rapeseed (Brassica campestris L.) has been purified 25-fold by fractionation with acetone and calcium phosphate gel. The purified enzyme was relatively stable when stored frozen at neutral pH, and was active over a broad range of pH, the optimum being at about pH 8.6 – 10.0. The enzyme required Mg++ and exhibited maximum activity only when the Mg++: ATP ratio was 1:1. Phosphorylcholine, ADP, and Mn++ inhibited the activity. In addition to choline, the enzyme preparation phosphorylated dimethyl- and diethyl-aminoethanol but not ethanolamine. The choline kinase from rapeseed is similar to that from yeast except that the former does not require sulphydryl groups for activation.

scholarly journals BRAIN MITOCHONDRIA

1963 ◽  
Vol 19 (2) ◽  
pp. 309-316 ◽  
Author(s):  
Diana S. Beattie ◽  
Howard R. Sloan ◽  
R. E. Basford
Keyword(s):  
High Speed ◽  
Brain Cortex ◽  
Lactate Production ◽  
Mitochondrial Fraction ◽  
Brain Mitochondria ◽  
Glycolytic Enzymes ◽  
Supernatant Fraction ◽  
Glycolytic Activity ◽  
High Speed Supernatant ◽  
Stimulation Of

A mitochondrial fraction prepared from calf brain cortex possessed negligible glycolytic activity in the absence of the enzymes of the high speed supernatant fraction. When mitochondria were added to a supernatant system supplemented with optimal amounts of crystalline hexokinase, a 20 per cent stimulation of glycolysis was observed. The supernatant fraction produced minimal amounts of lactate in the absence of exogenous hexokinase; the addition of mitochondria doubled the lactate production. The substitution of glycolytic intermediates for glucose as substrates as well as the addition of exogenous glycolytic enzymes to the supernatant fraction or supernatant fraction plus mitochondria indicated that the mitochondria contributed mainly hexokinase and phosphofructokinase. By direct assay of all of the enzymes of the glycolytic pathway, only hexokinase and phosphofructokinase were shown to be concentrated in the mitochondrial fraction. All other glycolytic enzymes were found to exhibit higher total and specific activities in the supernatant fraction.

CHOLINE KINASE OF RAPESEED (BRASSIGA CAMPESTRIS L.)

1957 ◽  
Vol 35 (10) ◽  
pp. 853-863 ◽  
Author(s):  
T. Ramasarma ◽  
L. R. Wetter
Keyword(s):  
Calcium Phosphate ◽  
Enzyme Preparation ◽  
Brassica Campestris ◽  
Maximum Activity ◽  
Choline Kinase ◽  
Aqueous Extracts ◽  
Neutral Ph ◽  
Sulphydryl Groups

Choline kinase from aqueous extracts of Polish rapeseed (Brassica campestris L.) has been purified 25-fold by fractionation with acetone and calcium phosphate gel. The purified enzyme was relatively stable when stored frozen at neutral pH, and was active over a broad range of pH, the optimum being at about pH 8.6 – 10.0. The enzyme required Mg++ and exhibited maximum activity only when the Mg++: ATP ratio was 1:1. Phosphorylcholine, ADP, and Mn++ inhibited the activity. In addition to choline, the enzyme preparation phosphorylated dimethyl- and diethyl-aminoethanol but not ethanolamine. The choline kinase from rapeseed is similar to that from yeast except that the former does not require sulphydryl groups for activation.

scholarly journals GENETIC ANALYSIS OF MUTANTS WITH A REDUCED Ca2+-DEPENDENT K+ CURRENT IN PARAMECIUM TETRAURELIA

Genetics ◽  
1985 ◽  
Vol 111 (3) ◽  
pp. 433-445
Author(s):  
Robert D Hinrichsen ◽  
Ed Amberger ◽  
Yoshiro Saimi ◽  
Anthony Burgess-Cassler ◽  
Ching Kung
Keyword(s):  
Genetic Analysis ◽  
High Speed ◽  
Behavioral Responses ◽  
Supernatant Fraction ◽  
Paramecium Tetraurelia ◽  
Wild Type ◽  
Wild Type Phenotype ◽  
K Current ◽  
High Speed Supernatant ◽  
K Currents

ABSTRACT Two mutants of Paramecium tetraurelia with greatly reduced Ca2+-dependent K+ currents have been isolated and genetically analyzed. These mutants, designated pantophobiac, give much stronger behavioral responses to all stimuli than do wild-type cells. Under voltage clamp, the Ca2+-dependent K+ current is almost completely eliminated in these mutants, whereas the Ca2+ current is normal. The two mutants, pntA and pntB, are recessive and unlinked to each other. pntA is not allelic to several other ion-channel mutants of P. tetraurelia. The microinjection of a high-speed supernatant fraction of wild-type cytoplasm into either pantophobiac mutant caused a temporary restoration to the wild-type phenotype.

Lipid synthesis in Lipomyces lipofer

1968 ◽  
Vol 46 (4) ◽  
pp. 303-313 ◽  
Author(s):  
F. A. McElroy ◽  
H. B. Stewart
Keyword(s):  
High Speed ◽  
Lipid Synthesis ◽  
Acid Synthesis ◽  
Mitochondrial Fraction ◽  
Maximum Activity ◽  
Rate Limiting Step ◽  
Cell Dispersion ◽  
Lag Period ◽  
Lipomyces Lipofer ◽  
High Speed Supernatant

The synthesis of lipids by cell-free fractions of Lipomyces lipofer was investigated by measuring the incorporation of acetate-14C and other radioactive precursors into lipid-soluble material. On the basis of cofactor requirements, the reaction to inhibitors, and the ability to incorporate appreciable quantities of malonate-14C, it was concluded that the malonyl-CoA pathway was the principal route of fatty acid synthesis in this yeast. By employing a means of cell dispersion less rigorous than previously used with yeasts, it was shown that the soluble portion (high-speed supernatant) of the cell was primarily involved in lipid synthesis. The mitochondrial fraction and the high-speed pellet (96 000 × g, 1 h) were inactive although the latter fraction was required for the maximum activity of the high-speed supernatant. The stimulatory activity of this pellet was very labile and showed a lag period before it became effective. Several lines of evidence suggested that the carboxylation of acetyl-CoA is the rate-limiting step in lipid synthesis which is accelerated by the high-speed pellet.

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