Growth and hemolymph trehalose levels of Heliothis zea larvae fed diets containing various sugars

• 1.1. The amount of sugar required for growth of Heliothis zea larvae on a chemically defined diet was determined.
• 2.2. Larvae grew well on fructose, galactose, sucrose, trehalose and raffinose diets but not on diets containing more than 0.5% glucose.
• 3.3. A starch diet did not promote rapid larval growth.
• 4.4. Hemolymph trehalose levels in 12-day-old larvae ranged from none to 45μmoles/ml.
• 5.5. A method for analysis of hemolymph trehalose by gas chromatography is described.

     

Nucleoside monophosphoramidate hydrolase from rat liver: Purification and characterization

• 1.1. Adenosine 5'-phosphoramidate hydrolase of 29 kDa was isolated from rat liver cytosol.
• 2.2. It consisted of two subunits of 14 kDa.
• 3.3. It hydrolyzed nucleoside 5'-monophosphoramidates into nucleoside 5'-monophosphates and ammonia, while it did not hydrolyze adenylyl phosphoramidate, adenylyl imidodiphosphate and N-phosphorylated compounds like phosphocreatine, N^ω-phosphoarginine, 6-phospholysine and 3-phosphohistidine.
• 4.4. Divalent cations and cyclic AMP had no effect on the hydrolytic activity.

     

Phosphofructokinase from the anterior byssus retractor muscle of Mytilvs edulis: Modification of the enzyme in anoxia and by endogenous protein kinases

• 1.1. Anoxia exposure resulted in a stable modification of the kinetic properties of 6-phosphofructo-1-kinase (PFK) from the anterior byssus retractor muscle (ABRM) of the sea mussel Mytilus edulis L.
• 2.2. Compared to the aerobic enzyme, the anoxic form of PFK. showed a reduced affinity for both substrates, fructose-6-phosphate (F6P) and ATP, and an increased sensitivity to inhibition by phosphoenolpyruvate.
• 3.3. To analyze the involvement of protein kinases in the modification of PFK, extracts from aerobic or anoxic muscle were incubated with ATP and Mg²⁺ plus protein kinase second messengers cyclic 3',5'-adenosine monophosphate (cAMP), cyclic 3',5'-guanosine monophosphate (cGMP) or Ca²⁺ plus phorbol 12-myristate 13-acetate (PMA).
• 4.4. Both forms of the enzyme responded to the presence of cAMP with a strong increase in affinity for F6P.
• 5.5. In response to cGMP affinity of the aerobic enzyme for F6P decreased whereas that of the anoxic enzyme form was not affected (at 0.5 mM ATP) or increased (at 3 mM ATP).
• 6.6. Incubation with Ca²⁺ + PMA had only a limited effect on PFK kinetics but appeared to enhance the response to cGMP when the three compounds were given together.
• 7.7. Treatment of PFK-aerobic with alkaline phosphatase resulted in a strong decrease in enzyme activity and affinity for F6P; subsequent treatment with cAMP reversed the effect on S_0.5 F6P.
• 8.8. The data indicate that PFK activity is altered during the aerobic-anaerobic transition by a change in the phosphorylation state of the enzyme and that cAMP and cGMP act oppositely to regulate PFK activity, and thereby alter glycolytic rate, during this transition.

     

The purification of kallikrein from human whole saliva

• 1.1. A quick and simple procedure is described for purifying kallikrein from human whole saliva. The enzyme has been purified about 2700-fold with a yield of approx. 30%.
• 2.2. The procedure is based on the immediate fractionation of saliva by ion exchange chromatography. This is followed by a combination of affinity and high performance liquid chromatography.
• 3.3. The results indicate that another protein component binds to the enzyme at pH 8.0.
• 4.4. The homogeneity of the enzyme has been demonstrated by gel electrophoresis in the absence as well as in the presence of sodium dodecylsulfate.
• 5.5. A mol. wt of 40,100±1800 has been calculated from gel electrophores is experiments.
• 6.6. Sedimentation equilibrium in an analytical ultracentrifuge gave a mol. wt of 39,700.
• 7.7. The amino acid composition has been determined and it confirms that the enzyme has a low isoelectric point.
• 8.8. The presence of tryptophan has been demonstrated by absorption and fluorescence spectroscopy.

     

Chemical structures of mono-, di-, tri- and tetraglycosyl glycerides in rice bran

• 1.1. Monoglycosyl monoglyceride, mono-, di-, tri- and tetraglycosyl diglycerides were isolated from rice bran and characterized for their chemical structures.
• 2.2. Monoglycosyl monoglycerides were characterized as Gal(β1' → 3)-1- or 2-monoacyl-sn-glycerol and Glc(β1' → 3)-1- or 2-monoacyl-sn-glycerol.
• 3.3. The structures ofmonoglycosyl diglyceride were Gal(β1' → 3)-1,2-diacyl-sn-glycerol and Glc(β1' → 3)-1,2-diacyl-sn-glycerol. Epimeric separation of the galactosyl and glucosyl glycerides was for the first time achieved by thin-layer chromatography.
• 4.4. The main diglycosyl diglyceride was shown to be Gal(α1' → 6')-Gal(β1' → 3)-1,2-diacyl-sn-glycerol.
• 5.5. The major structure of triglycosyl diglyceride was characterized as Gal(α1″' → 6″)-Gal(α1″ → 6')-Gal(β1' → 3)-1,2-diacyl-sn-glycerol.
• 6.6. The representative structure of tetraglycosyl diglyceride was for the first time established as Gal(α1″ → 6″')-Gal(α1″' α 6″)-Gal(α1″ → 6')-Gal(βl' → 3)-1, 2-diacyl-sn-glycerol.

     

Amino acid sequences around exofacial proteolytic cleavage sites of band 3 from bovine and porcine erythrocytes

• 1.1. Amino acid sequences of bovine and porcine band 3, an erythrocyte anion transporter, were determined.
• 2.2. The sequence of bovine band 3 was positioned to residues 519–599 (the numbering is based on human band 3), in which probably 6 residues were unidentified.
• 3.3. Binding site of DIDS (4,4'-diisothiocyanostilbene-2,2'-disulfonate), a potent anion transport inhibitor, was identified as Lys-539 in the bovine case.
• 4.4. A loop (residues 551–567), which provides exofacial proteolytic cleavage sites, contains only 53% homology between human and bovine, whereas the residues flanking it on either side are >84% homologous.
• 5.5. Furthermore, the loop of porcine band 3 was indicated to consist of a 6 or 7-residues short peptide as compared with those of other species.

     

Intracellular localization and characterization of 3-hydroxykynureninase in human liver

• 1.1. 3-hydroxykynureninase in human liver was present in cytosol and mitoehondria.
• 2.2. The cytosolic enzyme and mitochondrial enzyme had the same physiological and enzymic properties.
• 3.3. The enzyme had a mol. wt of 130,000 by gel filtration and isoelectric point of pH 5.9.
• 4.4. The enzyme was active for 3-hydroxykynurenine and kynurenine, and its activity ratio was 15:1. The apparent K_m values of the enzyme were 7.7 × 10⁻⁵M for 3-hydroxykynurenine, 1.0×10⁻³M for kynurenine and 2.5 × 10⁻⁶M for pyridoxal 5'-phosphate with 3-hydroxykynurenine.
• 5.5. Some other properties of purified enzymes are described.

     

Lysosomal acid deoxyribonuclease from vervet monkey livers—II: Enzymic properties

• 1.1. Primate liver lysosomal acid DNase is an endonucleolytic enzyme.
• 2.2. The enzyme has both 3'- and 5'-nucleotidohydrolase activities.
• 3.3. The oligonucleotides produced by DNase are polymers mainly about 30 mononucleotides long.
• 4.4. The Arrhenius plot shows a discontinuity with a transition temperature at 47°C, with an activation energy of 107 kJ/mol below and 67 kJ/mol above this temperature.
• 5.5. The activation enthalpy is 104kJ/mol and the entropy −0.498 kJ/mol/K.
• 6.6. The enzyme is subject to substrate inhibition and the K_m value is 159 × 10⁻³mM DNA-P.

     

Evidence that the 5' end of rabbit globin mRNA is hybridized with its 3' end

• 1.1. A radiopolyadenylated rabbit globin mRNA was treated with different concentrations of ribonuclease V₁ from cobra venom.
• 2.2. The enzymatic digests were chromatographed on an aminophenylboronate-agarose column, which specifically captured the cap structure i.e. n⁷G(5') ppp (5') NmP.
• 3.3. When the capture fragment was chromatographed on a Sephadex G-100 column, its size was smaller than the native molecule and also bore radioactivity, i.e. a poly(A) tail.
• 4.4. These results provide evidence that the 5' end (which encompasses the cap structure) of rabbit globin mRNA is hybridized and in close proximity to its 3' end.
• 5.5. We conclude that this conformation is required for messenger translation efficiency.

     

Structural features of carbohydrate chains in human salivary mucins

• 1.1. The structure of carbohydrate chains in the low and high molecular weight mucus glycoprotein forms from submandibular-sublingual saliva of individuals with blood group B was investigated.
• 2.2. Alkaline borohydride reductive cleavage of the glycoproteins yielded in each case a population of neutral (55%) and acidic (45%) oligosaccharide alditols ranging in size from 3 to 16 sugar units.
• 3.3. The predominant neutral oligosaccharides in both glycoprotein forms consisted of 16 and 15 sugar units arranged in triantennary fashion, and carried blood group B and I antigenic determinants.
• 4.4. Three of the oligosaccharides in each glycoprotein contained sialic acid and ranged in size from 3 to 12 sugar units. In two oligosaccharides sialic acid was linked to C3 of galactose and in one to C6 of N-acetylgalactosamine. The sulfated oligosaccharide in both glycoproteins was identified as a pentasaccharide with the sulfate ester group at C6 of N-acetylglucosamine.
• 5.5. The results demonstrate that contrary to the earlier view the low and high molecular weight mucus glycoprotein forms of human saliva contain identical carbohydrate chains.

     

Purification of the flavin-containing monooxygenase from mouse and pig liver microsomes

• 1.1. The microsomal flavin-containing monooxygenase has been purified from mouse and pig liver utilizing Cibacron-Blue Sepharose, Procion-Red agarose, and 2'5'-ADP Sepharose.
• 2.2. The enzymes had a final specific activity of 1200 and 954 nmol/min/mg protein from mouse and pig liver respectively.
• 3.3. The enzyme from both mouse and pig liver displayed typical flavoprotein spectra and appeared homogeneous by denaturing polyacrylamide gel electrophoresis.

     

Partial characterization of the hemoglobin from Hubbs' beaked whale (Mesoplodon carlhubbsi)

• 1.1. Blood samples were obtained from an adult female Hubbs' beaked whale and her fetus.
• 2.2. Two major hemoglobins were demonstrated by cellulose acetate electrophoresis and were purified by ion exchange chromatography from each specimen.
• 3.3. The relative amounts of these components were different in the adult and fetus.
• 4.4. Both of these hemoglobins have a higher affinity for oxygen than normal human hemoglobin.
• 5.5. Maternal and fetal hemoglobins were separated and the N-terminal amino acid of each of these hemoglobins was found to be valine.
• 6.6. Tryptic peptide separation and amino acid analyses of the purified polypeptide chains indicated that the hemoglobins of the fetal sample were identical to those of the maternal.

     

Identification of essential amino acids in the active center of thyroidal NAD+ glycohydrolase

• 1.1. Purified thyroidal NAD⁺ glycohydrolase has been subjected to the action of a number of group specific reagents in order to gain information concerning its mode of action.
• 2.2. Modification of histidyl residues with diethylpyrocarbonate strongly suppresses the NAD⁺ glycohydrolase activity. Inactivation with this reagent can be reversed to some extent by subsequent treatment with hydroxylamine.
• 3.3. NAD⁺ and ADP-ribose partially protect against inactivation with similar efficiencies.
• 4.4. The incomplete reactivation with hydroxylamine after diethylpyrocarbonate treatment and the selective inactivation by 2,4-pentanedione indicates that apart from one or more essential histidyl residue(s) also lysyl residues are important for activity. NAD⁺ and to a smaller extent ADP-ribose again protect against inactivation by 2,4-pentanedione.
• 5.5. The sensitivity of the enzyme towards N-ethyl-5-phenyl-isooxazolium-3'-sulfonate further points to the importance of carboxylate containing side chains.
• 6.6. The mechanistic implications of these results are discussed.

     

Influence of environmental factors on the deep and skin temperature in the European bison,Bison bonasus (L.)

• 1.1. In 43 European bison divided into three groups (Group A, 3–8-month-old calves; Group B, 18-month-7-year-old young bison; Group C, 12–24-year-old bison) the rectal, humerus region and abdomen region temperatures were measured.
• 2.2. The experiments were carried out in winter months, from mid-December to mid-March.
• 3.3. The mean rectal temperatures changed from 38.55°C in calves to 38.15°C in the oldest bison.
• 4.4. The mean temperatures of the humerus region changed from 20.69°C in calves to 21.49°C in older bison.
• 5.5. The mean temperatures of the abdomen region changed from 20.79°C in calves to 22.17°C in older bison (Gr. B).
• 6.6. The cluster analysis divided the bison into four groups named hot, warm, cool and cold bison.
• 7.7. Only air temperature measured 2 m above the ground and snow cover influenced the integrated bison temperature. Age, sex and mass as well as some environmental factors had no influence.
• 8.8. Measurements made 1 to nearly 4hr after a bison's death showed a drop in rectal temperature and mostly increases in temperatures of the humerus and abdomen regions.

     

Characterization of an acid phosphatase—Glycoprotein from barley seeds

• 1.1. An acid phosphatase from barley seeds germinated for 120 hr, that is not affected by the presence of cycloheximide, is a glycoprotein.
• 2.2. The enzyme has an optimum pH 4.9 and a mol. wt of 57,000 as determined by SDS-gel electrophesis.
• 3.3. The main sugar is galactose amounting to one half of the protein and it is as either mono or disaccharide.
• 4.4. Serine and glycine predominate and hydroxyproline is absent.

     

Increased muscle glucose uptake in response to chronic glyburide treatment is not related to changes in glucose transporter (GLUT4) protein

• 1.1. The mechanism of action of glyburide (a sulfonylurea) on muscle has been investigated by measuring glucose uptake and glucose transporter (GLUT4) protein levels after chronic glyburide treatment.
• 2.2. A dietary induced insulin resistant rat model (4 wk of high-fat, high-sucrose feeding) was given glyburide (2mg/kg/day) for 10 days and glucose uptake was measured in a perfused hindquarter preparation.
• 3.3. Protein levels of the GLUT4 glucose transporter were determined by Western analysis.
• 4.4. After 7 days of treatment, rats fed glyburide had lower blood glucose concentrations 2 hr (72 ± 5 vs 103 ± 12 mg/dl) and 24 hr (97 ± 7 vs 123 ± 7 mg/dl) after glyburide administration with no difference in serum insulin levels compared to vehicle treated animals.
• 5.5. Glucose uptake was approx doubled in basal state (0 insulin) in response to glyburide (2.8 + 0.4 vs 1.7 ± 0.2μ mol/g per hr).
• 6.6. Maximal insulin (100 nM) stimulated glucose uptake tended to be higher in the glyburide treated group, but did not reach statistical significance (8.0 ± 0.7 vs 7.0 ± 0.6 μmol/g per hr).
• 7.7. Western analysis revealed no significant effect of glyburide on the GLUT4 protein level in skeletal muscle.
• 8.8. These results suggest that glyburide alters glucose uptake through some mechanism other than alterations in the level of the GLUT4 glucose transporter protein.

     

Tetanus responses under rapid bath solution change: Electrotonic depolarization of transverse tubules may release Ca2+ from sarcoplasmic reticulum of Rana japonica skeletal muscle

• 1.1. Single skeletal muscle fibers were transferred from a normal Ringer solution to Na⁺ ion free solution, and vice versa, and tetanus responses were recorded immediately after the transfer.
• 2.2. Fractional tetanus tension recorded immediately after the displacement from the Na⁺ ion free solution to normal Ringer solution was dependent on fiber diameter.
• 3.3. Diffusion of Na⁺ ions along the transverse tubules was simulated [apparent diffusion constant was 3.11 × 10⁻⁶ (cm²/s)].
• 4.4. Our results suggest that the electrotonic spreading of membrane potential, caused by an action potential in the transverse tubules, could release Ca²⁺ ions from sarcoplasmic reticulum.

     

Distribution of beta-adrenergic binding in fractionated porcine adipocytes

• 1.1. The subcellular distribution of the porcine adipocyte beta-adrenergic receptor was studied in fractionated adipocytes.
• 2.2. The 30,000 g pellet obtained from hypotonically lysed cells contained membrane vesicles and mitochondria; it yielded approx 200–300 fmol dihydroalprenolol-bound receptors/mg protein.
• 3.3. Activity was increased to about 1000 fmol/mg protein after isolation of a plasma membrane fraction on a Percoll gradient.
• 4.4. The 5'-nucleotidase, succinate dehydrogenase and lactate dehydrogenase activities were usually enriched in compartments different from the ligand-binding activity.
• 5.5. Activity of porcine adipocyte 5'-nucleotidase, a purported plasma membrane marker enzyme, was not distributed in the same manner as the beta-adrenergic receptor.

     

Changes in Synaptic Proteins Precede Neurodegeneration Markers in Preclinical Alzheimer's Disease Cerebrospinal Fluid

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Highlights

• •Proteomic analysis of cerebrospinal fluid and identification of synaptic component.
• •Use of super resolution microscopy to verify synapse-specificity in human tissue.
• •Selective reaction monitoring MS (SRM) of synaptic panel in 3 cohorts of Alzheimer's disease cerebrospinal fluid.
• •Synaptic protein changes precede tau in preclinical Alzheimer's disease.

     

Properties of sweat,burro (Equus asinus) and man

• 1.1. Sweat of two burros (Equus asinus) was collected after two desert walks, one at 38°C, one at 41°C.
• 2.2. Sweat from one burro was about twice as concentrated as from the other. In that respect and in respect to concentrations of chloride, sodium and potassium their sweat was like that of man.
• 3.3. Low concentrations of bicarbonate were present in burros' sweat contrasted with little if any in man's.
• 4.4. Urea nitrogen plus ammonia nitrogen were found in higher concentrations in burros' sweat than in man's sweat.