The effect of different krill meals fed to laboratory rats on their blood indices.

1. In 102 laboratory rats fed with (a) the krill standardized meal, (b) the krill meal with low chitin content, (c) the casein diet with D,L-methionine, (d) the casein diet with D,L-methionine supplemented with the krill carapace meal, (e) the casein diet with D,L-methionine supplemented with ash from the krill standardized meal and (f) the control diet--"Murigran" standard pelleted feed; the different blood indices were investigated. 2. The mean values of following indices: the number of erythrocytes and leucocytes, the percentage of leucocytes, the corpuscular haemoglobin concentration and red blood cell diameter were similar in all experimental and control groups. 3. The mean values of haematocrit and haemoglobin levels, the mean corpuscular thickness and volume were lower in rats fed with the casein diet with D,L-methionine supplemented with the krill carapace meal than in other groups. 4. All kinds of investigated indices were similar in rats fed with krill meal with low chitin contents, whose parents received the standardized krill meal and no sex differences have been shown here.     

CTP-dependent lipid kinases of yeast

Membrane fractions from yeast Saccharomyces cerevisiae catalyzed a transfer of gamma-phosphate from [gamma-32P]CTP into membranous lipids. Phosphorylated compounds were identified as phosphatidic acid and dolichyl phosphate (DolP). The membrane fraction also catalyzed phosphorylation of the exogenous dolichol. The activity of the phosphorylating enzymes could be modified by the yeast growing conditions; i.e., the enzyme from yeast grown aerobically favored the synthesis of phosphatidate over dolichyl phosphate in the ratio of 3:1, whereas the membrane fraction from anaerobically grown yeast synthesized PA and DolP in the ratio of 0.5:1. The activity of the phosphorylating enzymes could also be modified by divalent cations and the concentration of detergents. Phosphorylation of lipids does not occur in the presence of [gamma-32P]ATP and is not influenced by the presence of UTP or GTP. This result points to the specific role of CTP as a gamma-phosphate donor for the synthesis of phosphatidate and dolichyl phosphates in the yeast system.     

Non-protein-mediated transfer of phosphatidic acid between microsomal and mitochondrial membranes

The transfer of phosphatidic acid between rat liver microsomes loaded with [32P]-phosphatidic acid and rat liver mitochondria was studied in the absence of added lipid transfer proteins. It was found that during 1 h at 37 degrees C in the medium containing 100 mM KCl, 20-30% of phosphatidic acid but only 2.5% of phosphatidylcholine were transferred. This spontaneous transfer of phosphatidic acid remained the same after pretreatment of microsomes and mitochondria with 125 mM KCl or microsomes alone with 1 mM Tris, pH 8.6, procedures reported to remove adsorbed lipid transfer proteins. This transfer was insensitive to thiol-blocking reagents. The initial rate of this non-protein-mediated transfer of phosphatidic acid was virtually independent of the concentration of the acceptor membranes (mitochondria), thus indicating that it occurs by diffusion of the phospholipid through the aqueous phase rather than by membrane collision. About 80% of phosphatidic acid synthesized in the outer mitochondrial membrane was recovered in the inner membrane after a 1-h incubation, pointing to a high rate of the intermembrane transfer of this phospholipid within intact mitochondrion.     

The use of a resin adsorbent to isolate cytokinins from sea water

XAD-4 resin was used to isolate cytokinins present in Baltic sea water. Four compounds were tentatively identified as 6-(3-methylbut-2-enylamino) purine and zeatin, and their ribosides.     

Amino acid neurotransmitters in the CNS. Characteristics of the acidic amino acid exchange

D-Aspartate exchange, defined as amino acid-stimulated D-[3H]aspartate efflux, was investigated in a preparation of rat brain synaptosomes. The efflux of radiolabelled D-aspartate was found to be enhanced by micromolar concentrations of externally added D- and L-aspartate, L-glutamate, L-cysteate and L-cysteinesulphinate. The stimulation of release by external amino acids followed Michaelis-Menten kinetics; the apparent Km values (in microM) were: 14.65 +/- 0.98 for D-aspartate; 8.00 +/- 1.5 for L-aspartate; 22.31 +/- 1.62 for L-glutamate; 6.76 +/- 0.3 for L-cysteate and 7.89 +/- 1.23 for L-cysteinesulphinate. The Vmax values for efflux were 2.16-4.06 nmol/min per mg protein. The exchange process was found to require external NaCl but was very little affected by increase in the external [K+]. The demonstration of exchange as a part of the transport process provides support for the suggestion that in synaptosomal preparations a substantial portion of influx and efflux of amino acid neurotransmitters occurs via a reversible membrane carrier.     

Isolation and properties of glycerol-3-phosphate oxidoreductase from human placenta

Glycerol-3-phosphate oxidoreductase (sn-glycerol 3-phosphate: NAD+ 2-oxidoreductase, EC 1.1.1.8) from human placenta has been purified by chromatography on 2,4,6-trinitrobenzenehexamethylenediamine-Sepharose, DEAE-Sephadex A-50 and 5'-AMP-Sepharose 4B approximately 15800-fold with an overall yield of about 19%. The final purified material displayed a specific activity of about 88 mumol NADH min-1 mg protein-1 and a single protein band on polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulphate. The native molecular mass, determined by Ultrogel AcA 44 filtration, was 62000 +/- 2000 whereas the subunit molecular mass, established on polyacrylamide gel in the presence of 0.1% sodium dodecyl sulphate, was 38000 +/- 500. The isoelectric point of the enzyme protein, determined by column isoelectric focusing, was found to be 5.29 +/- 0.09. The pH optimum of the placental enzyme was in the range 7.4-8.1 for dihydroxyacetone phosphate reduction and 8.7-9.2 for sn-glycerol 3-phosphate oxidation. The apparent Michaelis constants (Km) for dihydroxyacetone phosphate, NADH, sn-glycerol 3-phosphate and NAD+ were 26 microM, 5 microM, 143 microM and 36 microM respectively. The activity ratio of cytoplasmic glycerol-3-phosphate oxidoreductase to mitochondrial glycerol-3-phosphate dehydrogenase in human placental tissue was 1:2. The consumption of oxygen by human placental mitochondria incubated with the purified glycerol-3-phosphate oxidoreductase, NADH and dihydroxyacetone phosphate was similar to that observed in the presence of sn-glycerol 3-phosphate. The possible physiological role of glycerol-3-phosphate oxidoreductase in placental metabolism is discussed.     

The inhibitory effect of various fatty acids on aerobic glycolysis in Ehrlich ascites tumour cells

Inhibition of glycolysis in Ehrlich ascites tumour cells by saturated fatty acids, added either in form of potassium salts or incorporated into phosphatidylcholine liposomes, increases with the increasing carbon atom chain length and is independent of the concentration within the range of 0.1 to 1.0 mM. In contrast, the inhibition of glycolysis in the cytosolic fraction from Ehrlich ascites cells depends on the concentration of fatty acids. The content of ATP in Ehrlich ascites cells incubated with fatty acids increases with increasing carbon atom chain length, which leads to a crossing-over in the concentrations of pyruvate and 2-phosphoenolpyruvate. Lowering of the sum of both these metabolites by palmitate and stearate points to the inhibition not only of pyruvate kinase but also of other enzymes of early steps of glycolysis. Fatty acids in intact Ehrlich ascites cells inhibit all three key glycolytic enzymes but added to the cytosolic fraction affect mainly the activity of phosphofructokinase. The inhibition of pyruvate kinase by fatty acids is smaller in the cytosolic fraction from tumour cells than from liver and muscles.     

Glucuronides of monohydroxylated bile acids: specificity of microsomal glucuronyltransferase for the glucuronidation site, C-3 configuration, and side chain length

The ability of rat liver microsomes to catalyze UDP-glucuronic acid-dependent glucuronidation of monohydroxy-bile acids was examined. The following bile acids were used as substrates, each as the 3 alpha and 3 beta epimer: 3-hydroxy-5 beta-cholanoic acid (C24), 3-hydroxy-5 beta-norcholanoic acid (C23), 3-hydroxy-5 beta-bisnorcholanoic acid (C22), 3-hydroxy-5 beta-pregnan-21-oic acid (C21), and 3-hydroxy-5 beta-androstane-17 beta-carboxylic acid (C20). The corresponding glucuronides were chemically synthesized to serve as standards and were characterized by thin-layer and gas-liquid chromatography as well as by nuclear magnetic resonance. Enzymatic glucuronidation reactions were optimized with respect to pH for each product formed and the kinetic parameters for each reaction were measured. Analytical techniques necessary to separate products from unreacted substrates and to identify them included thin-layer chromatography, gas-liquid chromatography, and nuclear magnetic resonance. It was found that the 3 alpha epimers of the five bile acids listed above enzymatically formed 3-O-glucuronides, C24 being the best substrate, followed by C21 and C20; C22 and C23 gave rise to only small amounts of this product. The 3 beta epimers of all bile acids tested were poorer substrates, although by a factor that varied widely. In addition to the expected hydroxyl-linked glucuronide, three of the 3 alpha-bile acids (C23, C22, and C20) and at least one 3 beta-bile acid (C20), gave rise to a novel metabolite in which the 1-OH of glucuronic acid was esterified with the steroidal carboxyl group (carboxyl-linked glucuronide).     

The composition of liver chromatin proteins from embryos, chicks and adult chickens

The chemical composition of chromatin from the livers of 12-, 15- and 19-day-old embryos, of 1-day-old chicks and of adult chickens was analysed. The process of embryonic development is accompanied by an increase in non-histone chromatin proteins and chromatin RNA, as well as in the phosphorus content of chromatin phosphoproteins. The amount of these components decreases in the livers of 1-day-old chicks and adults. Two-dimensional polyacrylamide gel electrophoresis of acid-soluble chromatin proteins showed an increase in the amount of the H1 histone in 19-day-old embryos and adult chickens. Non-histone proteins of embryo liver chromatin showed a high content of the fraction of Mr of about 40 000; this was not the case for adult chickens. The non-histone protein fraction of Mr of about 120 000, characteristic of adult chicken liver proteins, was not found in the livers of 12- and 15-day-old embryos. Non-histone chromatin proteins isolated from the livers of animals of different age exhibited also quantitative differences.     

Immunological analysis of histone H2A from the slime mold Physarum polycephalum

Specific antibodies against the histone H2A from calf thymus were generated by injecting rabbits with complexes: histone H2A-RNA with a protein to RNA ratio of 3:1. In the microcomplement fixation assay the antibodies against the histone H2A from calf thymus immuno-reacted with the histone H2A from calf thymus but not with H2A from Physarum polycephalum. The histone H2A from calf thymus therefore appears to have an immunological determinant(s) which does not exist in H2A from Physarum polycephalum.