bluuebunny/crossref_metadata_2025 huggingface dataset:
Thermal Inactivation of Avian Liver and Muscle Fructose 1,6-Diphosphatases in the Presence of Fructose 1,6-Diphosphate, AMP, and Divalent Cations
Authors: Marquardt Ronald R. , Olson J. P.
Published: December 2009

The effects at various pH levels of fructose 1,6-diphosphate (FDP), AMP, CaCl2, MnCl2, and/or MgCl2 on the thermal stability of pure forms of avian liver and muscle fructose 1,6-diphosphatases were investigated. Both enzymes were more stable when stored at 20 °C than at 2 °C. Maximum stabilities of the two enzymes when incubated at various temperatures in a Tris buffer occurred at approximately pH 6. All of the above compounds when added to a pH 7.5 Tris buffer markedly increased the thermal stabilities of both enzymes. Individually, FDP provided the highest degree of protection for the muscle enzyme whereas MnCl2 was most effective with the liver enzyme; in all cases the degree of protection was concentration dependent. At pH 7.5 combinations of MgCl2, FDP, and AMP were most effective. In the presence of these compounds initial inactivation of the muscle and liver enzymes did not occur until respective temperatures of 74 and 78 °C (15 min) were reached; these compare with initial inactivation temperatures in the absence of these compounds of 55 and 59 °C. The interaction of MgCl2 and FDP with the two enzymes at varying pH levels showed markedly different patterns. At pH7.5 and 8.8 increasing levels of MgCl2 provided increasing levels of protection whereas at pH 6.2 increasing MgCl2 concentrations resulted in an enhanced degree of inactivation. In the presence of FDP an opposite pattern was observed at the various pH levels. The presence of phosphate in the buffer tended to yield results similar to those of FDP.
Role of Tocopherol in the Reduction of Mitochondrial NAD
Authors: Carabello Frank B.
Published: December 2009

Tocopherol deficiency had no effect on phosphate uptake linked to ascorbate–tetramethyl-p-phenylenediamine but did impair energy transfer associated with succinate-linked reduction of NAD. These results suggest that tocopherol deficiency uncouples site 1 but not site 3 related energy transfer.Marked differences were obtained with tocopherol deficiency and methyl linoleate hydroperoxide which showed that the effects of tocopherol deficiency on mitochondrial energy transfer could not be duplicated by in vitro addition of peroxides.Tocopherol administration via the portal vein, 45 min prior to extraction of tissue, reversed the effects of tocopherol deficiency. Intraportal diphenyl-p-phenylenediamine reversed lipid peroxidation but did not substitute for tocopherol in restoration of mitochondrial energy transfer. It was concluded that restoration of antioxidant activity does not assure restoration of energy transfer in tocopherol-deficient mitochondria.These results suggest that there is a specific requirement for tocopherol in succinate-linked reduction of NAD which would imply a cofactor function for tocopherol or a tocopherol-dependent metabolite. It was proposed that coenzyme Q could be a tocopherol-dependent metabolite which is required for energy transfer.
Some Characteristics of the Exchange of Lecithin between Rabbit Erythrocytes and Serum
Authors: Smith Norman , Rubinstein David
Published: December 2009

The exchange of lecithin between rabbit erythrocytes and serum has been investigated. The erythrocyte lecithin was labelled in vivo by administration of Na332PO4, which resulted in the acquisition of [32PJlecithin by the cells primarily by exchange, and in vitro by esterification with [3H]palmitate, [14C]linoleate, or [14C]linolenate. The rate of exchange of erythrocyte [32PJ-phospholipids was independent of specific serum lipoprotein fractions, but was related to the serum phospholipid concentration. Analysis of the erythrocyte inward and outward exchange rates of [32P]lecithin revealed that only 55% of the cellular lecithin pool takes part in the exchange. A similar conclusion can be reached by comparison of the fractional exchange rates of the acyl- and 32P-labelled erythrocyte lecithin, or by comparison of the ratio of acyl to 32P radioactivity of the erythrocyte lecithin with the ratio found in the serum after exchange. These data indicated that the proportion of the acyl-labelled erythrocyte lecithin pool (± S.E.M.) capable of exchange was 53 ± 5%, 64 ± 12%, and 95 ± 6%, when esterified with [14C]linoleate, [3H]palmitate, and [14C]linolenate, respectively. The differences in the proportion of the acyl-labelled erythrocyte lecithin pool undergoing exchange could not be attributed to the preferential exchange of a particular molecular species of lecithin, although the tetraenoic lecithin did exchange to a relatively greater degree. This suggests that cellular lecithin formed by esterification of linolenate is found only in the portion of the pool available for exchange, whereas that formed by esterification of linoleate and probably palmitate is distributed throughout the entire lecithin pool. The fractional exchange rate of [32P]lecithin of isolated erythrocyte membranes was 3 times that of the lecithin of intact cells. However, the difference in fractional exchange rates between [3H]palmityl-lecithin or [14C]linoleyl-lecithin and [32P]lecithin is reversed in the membranes. These observations suggest that a portion of the total cellular lecithin pool becomes available for exchange only upon disruption of the erythrocyte, and may be located in a relatively inaccessible portion of the membrane such as the inner surface.
Effect of Phosphatidylcholine on Triacylglycerol Synthesis in Rat Intestinal Mucosa
Authors: O'Doherty P. J. A. , Yousef I. M. , Kuksis A.
Published: December 2009

The acyltransferases involved in triacylglycerol synthesis were studied in mucosal microsomes from normal and bile fistula rats. Sustained biliary diversion (2–3 days) resulted in a 35–40% decrease in the activities of the mono- and diacylglycerol acyltransferases. In addition, the phospholipid/protein and phosphatidylcholine/phosphatidylethanolamine ratios were lower in the mucosal microsomes from the bile fistula rats. The addition of phosphatidylcholine and lysophosphatidylcholine to the mucosal microsomes resulted in restoration of the enzyme activities to control levels. Unsaturated species of phosphatidylcholine were more effective than saturated species. The phosphatides of ethanolamine, inositol, and serine were without effect. This suggested that the loss in enzyme activity was associated with a depletion of phosphatidylcholine in the mucosal microsomes. It is concluded that the mono- and diacylglycerol acyltransferases require phosphatidylcholine for optimal activity.
The Effects of Catecholamines, Glucagon, and Diet on Enzyme Activities in Brown Fat and Liver of the Rat
Authors: Hahn Peter , Kirby Lorne T.
Published: December 2009

The effects of a high carbohydrate diet fed to young rats for 24–48 h on phosphoenolpyruvate carboxykinase, malic enzyme, citrate-cleavage enzyme, and fatty acid synthetase could be overcome by injecting the animals with glucagon (1 mg/100 g body weight) or norepinephrine (20 μg/100 g body weight) four times a day. The same effect was achieved with 50 mg ephedrine added to the 24 h diet. The catecholamines were more effective in brown fat, whereas glucagon seemed somewhat more effective in the liver.
Multiple Forms of Potato Spindle-Tuber Metavirus Ribonucleic Acid
Authors: Singh R. P. , Michniewicz J. J. , Narang S. A.
Published: December 2009

Multiple infectious low-molecular weight ribonucleic acids from Scopolia sinensis plants, infected with potato spindle tuber metavirus were isolated by Bio-Gel P-200 gel filtration columns and the slowest-moving form (infectious peak III) was eluted with bulk of 4 S-RNA.
Changes in Seryl-tRNA Formative in Developing Chick Muscle
Authors: Nwagwu Mark , Lianga John
Published: December 2009

As a prelude to an analysis of the dependence of muscle protein synthesis on aminoacyl tRNA's, we have investigated the rates of seryl-tRNA formation, in vitro, by aminoacylating systems isolated from 11-, 14-, and 17-day chick embryonic muscle. The results show that the combination of 14-day tRNA and 14-day aminoacyl synthetase is the most efficient in seryl-tRNA formation. We have also studied the qualitative and quantitative changes in seryl-tRNA prepared from 11-, 14-, and 17-day embryonic chick muscle by chromatography of seryl-tRNA on benzoylated DEAE-cellulose columns. The results show that, although there are no qualitative differences in the chromatographic patterns of seryl-tRNA from the different ages, there are significant quantitative differences between the patterns for 11-day and 17-day seryl-tRNA on the one hand, and the pattern for 14-day seryl-tRNA on the other.
The Decay of Xenopus laevis Mitochondrial rRNA in the Presence of Ethidium Bromide or Actinomycin D
Authors: Young P. G. , Zimmerman A. M.
Published: December 2009

The effect of ethidium bromide and actinomycin D on prelabelled mitochondrial RNA was studied in Xenopus laevis embryos. Mitochondrial rRNA was found to decay with a half-life of 2–2.5 h following an initial lag of < 2 h. There was an initial increase in incorporation into mitochondrial rRNA in the presence of actinomycin D but not in the presence of ethidium bromide. Radioactivity in mitochondrial 4 S RNA rises and then is relatively stable for the duration of the chase period. Cytoplasmic rRNA and 4 S RNA are stable under the chase conditions used, and, in the presence of ethidium bromide, cytoplasmic RNA continues to be synthesized.
Controlled Cleayage of Phage R17 RNA within the Virion by Treatment with Ascorbate and Copper (II)
Authors: Wong K. , Morgan A. R. , Paranchych W.
Published: December 2009

A method is described for the degradation of R17 phage RNA in situ using vitamin C and copper (II). The procedure permits the controlled cleavage of the phage genome without affecting in any gross way the properties of the capsid proteins nor the secondary structure of the RNA. Losses of infectivity of the order of 6–7 logs were readily obtained without causing any detectable changes in the attachment capacity of phage nor in the structural integrity of the virion. Studies on the effect of oxygen, catalase, and superoxide dismutase on this reaction suggested that hydroxyl radicals generated from H2O2 may be the reactive species responsible for the cleavage of phage RNA.
Lipid and Temperature Dependence of Membrane-Bound ATPase Activity of Acholeplasma laidlawii
Authors: Hsung Jean-Cheui , Huang Leaf , Hoy Daniel J. , Haug Alfred
Published: December 2009

Arrhenius plots (15–45 °C) of the ATPase activity of Acholeplasma laidlawii membranes enriched with either arachidoyl or saturated short-chain groups exhibited a pronounced discontinuity in slope around 25–30 °C, absent in oleoyl-enriched membranes. The membrane lipid fluidity was measured with a stearic acid spin label. At the growth temperature (37 °C) the membrane lipid fluidity was identical for all three kinds of enriched membranes. For membranes enriched with saturated acyl chains, a plot of the electron paramagnetic resonance anisotropy parameter 2T11vs. the reciprocal of temperature showed a biphasic profile with a discontinuity in slope around 25–30 °C. For oleoyl-group enriched membranes this plot yields only a straight line between 15 and 50 °C. Thus, lipids do play a role in the functioning of the membrane-bound ATPase. This enzyme is probably localized in the more fluid regions of the membrane.
The Conversion of Cytochrome Oxidase to an Inactive Form of the Enzyme by Water Dialysis
Authors: Kornblatt Jack A. , Williams G. R.
Published: December 2009

A modification of beef-heart cytochrome oxidase that results in loss of catalytic capacity has been produced by extensive dialysis of the enzyme against water. The enzyme after dialysis retains all the heme, 80% of the phospholipid, and 60% of the copper originally associated with it. A comparison of the circular-dichroism (C.D.) spectra of the two forms has led to the conclusion that the heme of the enzyme is randomly oriented in the dialyzed form. Analysis of the rate at which this randomization occurs has shown that it is not the primary event occurring during dialysis. Prior to the loss of the C.D. spectrum of the dialyzed material there is a loss of enzymatic activity. Attempts to regenerate the original from the water-dialyzed enzyme by incubating the latter with a phospholipids–cholate mixture have been unsuccessful.
Enzyme Increase by Folate and Methotrexate in Cultured Mammalian Cells
Authors: Hillcoat B. L. , Marshall L.
Published: December 2009

Folate and methotrexate markedly increased the activity of dihydrofolate reductase in cultured cells of human origin. Folate did not have this effect in the cells of other species tested, whereas methotrexate increased enzyme activity in some other species. Neither compound increased enzyme activity in a somatic hybrid, although such an increase occurred in one of the parent cell lines. Active enzyme synthesis appeared necessary for the effect of folate to occur. Degradation of the increased enzyme after cycloheximide treatment of cells proceeded at least as rapidly as in control cultures and was associated with a decrease in the intracellular folate pool, whereas folate added after 48 h of culture stabilized the enzyme at this same intracellular concentration. Folate, added at 24 h of culture, produced the greatest increase in activity, but the subsequent rate of decrease was greater than that when folate was added at 48 h, although the intracellular concentration of folate in the latter case was only half that in the former.
Biological Activity of Ribose-Modified N6-(Δ2-Isopentenyl)adenosine Derivative
Authors: Chen C. M. , Smith O. C. , Hartnell G. F.
Published: December 2009

An analog of N6-(Δ2-isopentenyl)adenosine, 2-O-(1R-(9-N6-(Δ2-isopentenyI)adenyl)-2-hydroxyethyl) glycerol was prepared by cleavage of the 2′, 3′ carbon–carbon bond of the ribose moiety of N6-(Δ2-isopentenyl)adenosine by the technique of periodate oxidation and borohydride reduction of the dialdehyde to the diol. The net rate of uptake of this analog by tobacco-pith tissue cultures was about 40% of the N6-(Δ2-isopentenyl) [8-14C]-adenosine. The enzymatic hydrolysis of the glycosidic bond of this analog was not detected, but the N6-isopentenyl side chain was modified in the tobacco-pith tissue cultures. It was not a substrate for adenosine deaminase and not incorporated into tRNA. The relative biological activities in the tobacco-tissue culture bioassay suggested that the modification of the ribosyl moiety of N6-(Δ2-isopentenyl) adenosine reduced but did not abolish its ability to promote cell growth.
Phosphoribosyl Pyrophosphate Degradation in Human Tissues
Authors: Fox Irving H. , Marchant Pamela J.
Published: December 2009

Enzymatic hydrolysis of phosphoribosyl pyrophosphate (PP-ribose-P) in dialyzed human tissue homogenates had a specific activity ranging from 0.15 to 1.37 μmol mg−1 h−1. Our observations on human placenta have characterized this reaction: (1) PP-ribose-P was degraded in the absence of Mg with stoichiometric release of Pi at 37 °C; (2) the reaction occurred from pH 5 to 10.5 with the maximum activity in the alkaline range; (3) PP-ribose-P degrading activity was localized mainly in the microsomal fraction; (4) alkaline phosphatase rather than 5′-nucleotidase was responsible for the degradation of PP-ribose-P; (5) the Km for PP-ribose-P was 0.3 mM; (6) PP-ribose-P hydrolysis was altered by many phosphorylated compounds, both nucleoside and sugar derivatives, Pi, and PPi in concentrations ranging from 0.1 to 10.0 mM.
The Specificity of Oxidase and Kinase Preparations from Pseudomonas fluorescens towards Deoxyfluoromonosaccharides
Authors: Taylor N. F. , Hill L. , Elsenthal R.
Published: December 2009

With partially purified enzyme preparations from cell-free extracts of Pseudomonas fluorescens, 3-deoxy-3-fluoro-D-glucose and 3-deoxy-3-fluoro-D-gluconic acid are substrates for glucose oxidase (EC 1.1.3.4.) and gluconate dehydrogenase (EC 1.1.99.3), with Km values 18.2 mM and 11.8 mM, respectively. The same enzymes that oxidize glucose and gluconic acid probably oxidize 3-deoxy-3-fluoro-D-glucose and 3-deoxy-3-fluoro-D-gluconic acid. The latter fluorinated carbohydrates and the presumed formation of 3-deoxy-3-fluoro-2-keto-D-gluconic acid, which has been isolated as a calcium salt and characterized, are not substrates for gluconokinase (EC 2.7.1.12). Both 3-deoxy-3-fluoro-D-glucose and 3-deoxy-3-fluoro-D-gluconic acid act as competitive inhibitors of this enzyme preparation for gluconate, with Ki values 47.5 mM and 14.8 mM, respectively.
Studies of the Regulation of Purine Nucleotide Catabolism
Authors: Lomax Christopher A. , Bagnara Aldo S. , Henderson J. Frank
Published: December 2009

Ehrlich ascites tumor cells containing radioactive ATP were incubated in vitro with a range of concentrations of 2-deoxyglucose in order to produce different rates of ATP catabolism. Concentrations of all radioactive products of ATP catabolism were measured, and apparent rates of adenylate deaminase and inosinate dehydrogenase and of adenylate and inosinate dephosphorylation were calculated. It was concluded that these processes were regulated primarily by the rate of formation of substrate, and to a lesser extent in some cases, by substrate concentration. No evidence was obtained for regulation of these processes by the concentration of ATP. The deoxyglucose-induced catabolism of radioactive GTP was also studied. When ATP catabolism was induced by incubation with 2,4-dinitrophenol, time courses of accumulation of purine nucleoside monophosphates and rates of alternative pathways of their metabolism were quite different than when deoxyglucose was used.
The Demonstration of O-Acetylated Sialic Acids in Colonic Epithelial Glycoproteins
Authors: Reid P. E. , Culling C. F. A. , Tsang Wai-Chiu , Clay M. G. , Ramey C. W. , Dunn W. L.
Published: December 2009

Glycoproteins have been isolated from a 1 M sodium chloride extract of the colonic epithelial cells of man and rat by a combination of Agarose gel and DEAE-cellulose chromatography. The glycoproteins contain O-acetylated or O-esterified sialic acids of at least four types: (a) unsubstituted or possibly substituted at position C-9, (b) substituted at C-4 and possibly at C-9, (c) substituted at C-7 and/or C-8, and (d) substituted at C-4 as well as at C-7 and/or C-8.
Studies on Acute Phase Proteins of Rat Serum. V. Effect of Induced Inflammation on the Synthesis of Albumin and α1-Acid Glycoprotein by Liver Slices
Authors: Jamieson J. C. , Morrison K. E. , Molasky D. , Turchen B.
Published: December 2009

Liver slices from normal rats and those suffering from inflammation for 24–48 h were incubated with L-[14C]leucine or D-[I4C]glucosamine. Immunological techniques coupled with radioautography indicated that the microsome fraction prepared from slices contained the subcellular site of synthesis of the polypeptide chain of serum albumin, and the polypeptide and carbohydrate chains of α1-acid glycoprotein; both proteins were also present in the medium in labelled forms. The contents of albumin and α1-acid glycoprotein in the medium and in extracts of liver from experiments with liver slices from control rats and 8–72 h experimental rats were determined using the quantitative precipitin technique. There was a net increase in synthesis of both proteins when slices from control and experimental animals were used, the increase showing up in medium proteins. However, slices from livers from 8–72 h experimental rats had a greater capacity for synthesis of α1-acid glycoprotein and a lower capacity for synthesis of albumin than slices from livers from control rats, the greatest changes occurring with slices from 24 h experimental rats. Changes in synthetic capacities of liver slices from experimental rats for albumin and α1-acid glycoprotein were always accompanied by large increases in specific radioactivities of total medium proteins when experiments involved incubation of slices with L-[3H]leucine and D-[14C]glucosamine. It is suggested that the increase in specific radioactivities of medium proteins following incubation of liver slices from experimental rats with labelled leucine and glucosamine is a characteristic of the response of liver to inflammation, and reflects changes in the capacity of liver for the synthesis of α1-acid glycoprotein and other acute phase serum proteins.
Electrophoretic Characterization of Porcine Pancreatic (Pro)elastases A and B
Authors: Ledoux Michel , Lamy François
Published: December 2009

Two porcine pancreatic zymogens can be separated by free electrophoresis on a sucrose gradient. After activation by trypsin, both enzymes can hydrolyze completely the fibrous protein elastin. One of the two proteins, proelastase B, has, in addition, an esterolytic activity towards N-acetyl-L-tyrosine ethyl ester. The other, proelastase A, does not possess it. The activation products of the zymogens have been tagged with radioactive diisopropylfluorophosphonate and separated by polyacrylamide-gel electrophoresis. Proelastase A gives only one active species, pancreatopeptidase E, but three distinct proteins can be obtained from proelastase B. Elastases A and B exhibit an important synergism when acting together upon a purified elastin lacking microfibrils. Trypsin has considerably less synergistic activity, and chymotrypsin has practically none.
Effects of Changes in Environmental Lighting Upon Levels of Hydroxyindole O-Methyltransferase Activity in the Developing-Chick Pineal Gland
Authors: Wainwright S. D.
Published: December 2009

The level of hydroxyindole O-methyltransferase (HIOMT) activity in the pineal gland of developing chicks raised under constant illumination rose more rapidly and to higher values than in the gland of birds maintained in constant darkness. Rates of net increase in activity, and levels of activity attained, for birds raised under a diurnal cycle of illumination were intermediate between those maintained in constant light or darkness. Under each of the lighting conditions, the course of increase in enzymic activity was markedly affected by variations in an unidentified factor, the source of which appeared to be the hatching eggs.Birds transferred from constant light to the dark showed either an arrest of increase in enzyme activity or a loss of activity until the level equalled that observed for chicks of the same age raised in constant darkness. Chicks transferred from constant darkness to constant illumination showed marked increases in levels of enzyme activity at rates comparable with the maximal values observed with birds maintained under constant illumination, regardless of age and without delay. No diurnal cycle in level of HIOMT activity was observed in the pineals of 15-day birds.
The Control of Pyruvate Kinases of Escherichia coli. II. Effectors and Regulatory Properties of the Enzyme Activated by Ribose 5-Phosphate
Authors: Waygood E. Bruce , Rayman M. Khalil , Sanwal B. D.
Published: December 2009

The pyruvate kinases of Escherichia coli activated by ribose 5-phosphate (RP) has been partially purified. The active form of the enzyme has a molecular weight of about 180 000 as judged by sucrose density gradient centrifugations and Sephadex G-150 chromatography. On dissociation in the absence of sulfhydryl reagents such as dithiothreitol, the enzyme is inactivated and it has a molecular weight of about 110 000. Various substrates and effectors of the enzyme, with the exception of phosphate, do not influence the association–dissociation equilibrium of the enzyme.The enzyme, unlike pyruvate kinases from many other sources, is not activated by potassium ions. Sulfate and phosphate ions are inhibitory to the enzyme. Phosphate seems to be an allosteric inhibitor and its effect is completely antagonized by activators. The enzyme is activated in an allosteric manner by two classes of compounds, nucleoside monophosphates and sugar phosphates of the hexose monophosphate pathway. Amongst the nucleotides, guanosine 5′-phosphate and adenosine 5′-phosphate are the most effective activators. Amongst the hexose monophosphate pathway intermediates, RP is the most powerful activator, with apparent activation constants as low as 1 μM. Sugar phosphates esterified at C-1 or both terminal positions are entirely ineffective in activation. The effectors act by changing the Michaelis constant for the substrates. Both of the substrates of the enzyme, adenosine diphosphate and phosphoenolpyruvate, yield cooperative rate–concentration plots in the presence of unsaturating concentrations of the fixed changing substrate. The initial velocity plots for both substrates become hyperbolic in the presence of saturating concentrations of RP.
Characteristics of Action of Prostaglandins on Cyclic AMP Accumulation in Rat Anterior Pituitary Gland
Authors: Borgeat Pierre , Labrie Fernand , Garneau Pierre
Published: December 2009

Prostaglandins (PGs) were found to lead to a marked stimulation of cyclic AMP accumulation in rat anterior pituitary gland in vitro in the following decreasing order of potency: [Formula: see text]. The effect of PGs is potentiated by theophylline. The stimulatory effect of PGs on cyclic AMP accumulation is already detected 2 min after the addition of 1 × 10−7 to 1 × 10−6 M PG E2 and its maximal effect is reached after approximately 30 min of incubation, with a progressive decrease toward basal cyclic AMP levels at later time intervals. Increased intracellular cyclic AMP concentrations are accompanied by an increased release of the nucleotide into the incubation medium. Complete removal of Ca2+ from the incubation medium by addition of EGTA was found to increase the stimulatory effect of PG E2 on cyclic AMP accumulation. The action of PGs on hormonal release and cyclic AMP accumulation support the hypothesis of a role of PGs in the mechanism of anterior pituitary hormone (particularly growth hormone) release.
Carboxylesterases (EC 3.1.1). Amino Acid Composition of Liver Carboxylesterases
Authors: Scott Keith , Zerner Burt
Published: December 2009

The amino acid compositions of the carboxylesterases from chicken, horse, ox, sheep, and pig livers are reported and compared. As would be expected for this homologous series, the compositions show a general similarity. However, there are some significant differences, but the degree to which particular pairs of enzymes differ is consistent with the evolutionary history of the species from which they were isolated.
Photooxidation of Dinitrophenylhistidine-200 Human Carbonic Anhydrase B
Authors: Kandel M. , Gornall A. G. , Lam L. K. , Kandel S. I.
Published: December 2009

Partial inactivation of τ-dinitrophenylhistidine-200 human carbonic anhydrase B, induced by visible light, followed first order kinetics (kapp = 6.05 × 10−2 min−1). After 50 min the τ-dinitrophenylhistidine (τ-DNP-histidine) content decreased to a negligible level, but the illuminated enzyme retained, at pH 7.6, approximately 9.2% of the esterase activity of the native enzyme. The following lines of evidence suggest that the loss of activity results from the destruction of τ-DNP-histidine-200. (1) No significant loss of amino acid other than τ-DNP-histidine was detected after illumination. (2) The rate of loss of activity correlated well with the loss of τ-DNP-histidine. (3) In the photooxidized enzyme the DNP moiety was retained but had lost the characteristic sensitivity of τ-DNP-histidine to nucleophilic attack. Titration of the illuminated enzyme with acetazolamide indicated that the residual activity is an intrinsic property of the modified enzyme. The chromatographically purified photooxidized enzyme migrated as a single band on isoelectrofocusing in polyacylamide gel, and at pH 7.6 possessed 7.5% esterase activity relative to the native enzyme. By establishing effective destruction of histidine-200, it can be concluded that neither the π N nor, as previously shown, the τ N of histidine-200 is critical for the catalysis.
Thiocyanate Ion Formation in Rapeseed Meals
Authors: Srivastava V. K. , Hill D. C.
Published: December 2009

Meal prepared from unheated rapeseed (Brassica napus cv. Zephyr) showed the presence of thiocyanate ion, while meal from heated seed of the same cultivar did not show detectable amounts. Unheated seed meal on autolysis, and heated seed meal on incubation with thioglucosidase, yielded increased amounts of thiocyanate ion. Various commercial rapeseed meals showed the presence of thiocyanate ion only after enzyme incubation. Low glucosinolate, cv. Bronowski, and higher glucosinolate, cv. Zephyr on enzymic incubation yielded comparable amounts of thiocyanate ion, suggesting that the precursor responsible in the two varieties was the same and present in similar quantities. No formation of thiocyanate ion was observed on incubation of sinigrin with thioglucosidase. Rats dosed with heated meal, containing intact glucosinolate, showed a slight increase of thiocyanate ion in the urine as compared with control rats dosed with water, while a relatively large increase followed dosing with sinigrin. Rats dosed with meal containing free thiocyanate ion excreted the ingested thiocyanate ion almost quantitatively.
Effects of Intraventricular Injections of N6,O2′-Dibutyryl Adenosine 3′,5′-Monophosphate on the Rat Brain Cholinergic System
Authors: Askew William E. , Ho Beng T.
Published: December 2009

Intraventricular injections of dibutyryl cyclic AMP effected increases of acetylcholine in the cerebral cortex – striatal and subcortical regions. No increase in choline acetyltransferase activity was observed; however, there was a significant decrease in acetylcholinesterase activity in the subcortical region.
Isolation and Partial Characterization of a Sulfogalactoglycerolipid from Rat Brain
Authors: Levine M. , Kornblatt M. J. , Murray R. K.
Published: December 2009

The brains of adult rats were analyzed for the presence of 35SO4-containing glycolipids following intraventricular injection of Na235SO4. Radiochromatographic analyses revealed the presence of two minor 35SO4-containing glycolipids, in addition to sulfogalactosylceramide. One of these two minor sulfolipids was isolated and tentatively identified as a 1-O-alkyl-2-O-acyl-3-(3′-sulfogalactosyl)-glycerol, a compound recently demonstrated to be the major glycolipid of mammalian testis. The alkyl and acyl compositions of the compound from rat brain are more heterogeneous than those from rat testis. The non-sulfated form of the galactoglycerolipid was also detected in rat brain. The amount of the sulfogalactoglycerolipid in rat brain is 0.19 μmol per gram wet weight, approximately one-third of the amount in rat testis (per gram wet weight), and is approximately one-fifteenth that of sulfogalactosylceramide in rat brain. The possible significance of the common occurrence in brain and testis of sulfated and non-sulfated galactolipids is discussed.
Wheat Embryo Ribonucleates. V. Generation of N2-Dimethylgnanylate When 'Fully Sequenced' Homogeneous Species of Transfer RNA are Used as Substrates for Wheat Embryo Methyltransferases
Authors: Kwong T. C. , Lane B. G.
Published: December 2009

1. When S-adenosyl[methyl-14C]methiomne and various species of transfer RNA are used as substrates for wheat embryo methyltransferases, the principal site of guanylate-N2 methylation can be shown to be a G-residue between the stems of the dihydrouridine and anticodon loops. This common site of guanylate-N2 methylation is referred to as the interstem target site.2. When the interstem target site is the non-terminal G-residue in a G-C-G-C sequence, as in the cases of Escherichia coli tRNA1Leu, tRNAIle, and tRNA3Ser, there is preponderant dimethylation to yield N2-dimethylguanylate.3. When the interstem target site is part of a U-C-G-U sequence, as in the case of E. coli tRNAtMet, there is diminished dimethylation and correspondingly increased monomethylation to yield N2-monomethylguanylate.4. When the interstem target site is the non-terminal G-residue in an A-U-G-G sequence, as in the case of yeast tRNAAsp, there is negligible dimethylation and almost exclusive monomethylation to yield N2-monomethylguanylate.5. The concerted way in which the primary, secondary, and tertiary structures of tRNA molecules might influence the efficacy of these methylations is the subject of a brief discussion. Attention is also focused on the evolutionary and molecular basis for the generally non-random distributions of methylated oligonucleotide sequences in ribosomal and transfer ribonucleates.
Phosphatidylethanolamine in Liver Mitochondria and Endoplasmic Reticulum; Molecular Species Distribution and Turnover
Authors: Parkes J. G. , Thompson W.
Published: December 2009

Phosphatidylethanolamine from mitochondria and microsomes of guinea pig liver was separated by thin-layer chromatography into eight different classes differing in degree of unsaturation. The fatty acid compositions and molar proportions of each class isolated from microsomes were very similar to the corresponding class in mitochondria. In both organelles about half of the total was dienoic species while tetraenes comprised approximately 20%. Stearic acid was the major saturated fatty acid and in each membrane a greater selectivity for stearate over palmitate was found in each sub-class of phosphatidylethanolamine, when compared with the corresponding class of phosphatidylcholine.Following the intraperitoneal injection of [2-3H]glycerol, the labelling of each molecular class of phosphatidylethanolamine showed very similar progressions in microsomes and mitochondria over a 3 h interval. In both organelles the highest relative specific activity was attained by penta-plus hexaenoic classes, while the large dienoic class had the lowest relative activity, which, however, increased with time. Analysis of the dienoic class of phosphatidylethanolamine from whole liver showed it to be constituted by a rapidly turning over palmitoyl–linoleoyl fraction and a slowly labelled stearoyl–linoleoyl fraction, a pattern also exhibited by dienoic phosphatidylcholines.The similarities in profile of molecular classes of phosphatidylethanolamine and in the kinetics of labelling in vivo point to a close metabolic relation between the lipids of both organelles, suggestive of a transfer of different molecular classes at comparable rates from the endoplasmic reticulum, the site of synthesis, to the mitochondria. This is consistent with numerous other studies in vitro that have demonstrated inter-organelle exchange of lipids.
Kinetics and Reaction Mechanism of the Carbamylphosphate Synthetase of a Multienzyme Aggregate from Yeast
Authors: Aitken David M. , Lue Peter F. , Kaplan J. Gordin
Published: December 2009

We have studied the kinetics and reaction mechanism of the carbamylphosphate synthetase of an enzyme aggregate functioning in the pyrimidine pathway of yeast. Mg–ATP was found to be one of the substrates of the enzyme reaction which was activated by free Mg2+ and inhibited by free ATP. Feedback inhibition by UTP was non-competitive with respect to both glutamine and bicarbonate. Potassium ions were essential for activity and could not be replaced by sodium. Glutamine could be replaced partially by ammonium ions as nitrogen donor. A bicarbonate-dependent cleavage of ATP was shown to take place in the absence of L-glutamine; L-glutamate was a competitive inhibitor of L-glutamine and the enzyme was shown to synthesize ATP when incubated with ADP and carbamyl phosphate. The reaction mechanism was found to involve sequential addition of the substrates bicarbonate and Mg–ATP and release of ADP, followed by addition of the third substrate glutamine. The purine nucleotide XMP had a pronounced activating effect on the enzyme, acting at a site different from that of UTP. Saturating levels of Mg–ATP eliminated this activation.
Two Forms of a Mitochondrial Endonuclease in Neurospora crassa
Authors: Martin Charles E. , Wagner Robert P.
Published: December 2009

Mitochondrial nuclease activity in Neurospora crassa occurs in membrane-bound and soluble forms in approximately equal proportions. These activities apparently are due to the same enzyme, which has an approximate molecular weight of 120 000. A portion of the insoluble enzyme appears to be associated with the inner mitochondrial membrane and is resistant to solubilization by detergent treatment as well as by physical disruption methods.
Amino Acid and Protein Metabolism During Differentiation (Sclerotization) of the Myxomycete Physarum flavicomum
Authors: Henney Jr. Henry R. , Maxey Glenna
Published: December 2009

Protein synthesized by growing plasmodia of Physarum flavicomum was steadily degraded when the plasmodia were induced to differentiate (form sclerotia). Protein synthesis occurred during the initial one-fifth (9 h) of the 48 h differentiation period, but most of this protein was also degraded shortly after its synthesis. Amino acids were primary catabolites during the differentiation process, and catabolism was extensive, even in the presence of dextrose. Glutamic acid was catabolized at a rate about two and a half or three times greater, respectively, than that observed for valine and arginine. Active transport systems for amino acids appeared to be present and to remain functional in P. flavicomum during differentiation. Amino acids included in the sclerotization media were rapidly accumulated into the cell pool and protein fractions. Intracellular amino acids were actively retained and were not released into the medium during differentiation.Differentiation of this Myxomycete, therefore, is characterized by a change in the metabolism of the sclerotizing plasmodium to an autolytic type, as cellular proteins and amino acids are actively catabolized during the formation of the dormant sclerotia.
Reconstitution of 48 S Particles with Subunit Association Activity from Escherichia coli 50 S Ribosomal Components
Authors: Chu F. K. , Maeba P. Y.
Published: December 2009

Previous studies have shown that 50 S ribosomal proteins bind sequentially to RNA's of the 50 S subunit via temperature-dependent steps (0 → 37 → 55 °C) to form 28, 32, (43) and 48 S particles (Chu, F. K., and Maeba, P. Y. (1973) Can. J. Biochem. 51, 129–139). When three equivalents of 50 S ribosomal proteins are used per equivalent of 23 S RNA, the yield of reconstituted 48 S particles is greatly increased. The addition of 10 mM spermidine increases the rate of reconstitution and abolishes the need for prior incubation at lower temperatures when 48 S particles are formed at 55 °C. In the presence of excess 50 S ribosomal proteins and 10 mM spermidine, 48 S particles formed at 50 °C are able to associate with 30 S subunits in a stoichiometric manner to form 70 S complexes. These 48 S particles compete effectively with 50 S subunits for 30 S subunits and have an association activity equal to that of 50 S subunits. As compared to the 34 proteins in 50 S subunits, 4 are either absent or present in marginal amounts, 11 in reduced amounts, and 19 in approximately normal amounts in the 48 S particles.
Effect of Phenols on the Oxidation of Estradiol by Uterine Peroxidase
Authors: McNabb T. , Sproul J. , Jellinck P. H.
Published: December 2009

The effect of various phenols on the conversion of [4-14C]estradiol to water-soluble products by estrogen-induced uterine peroxidase (EC 1.11.1.7) has been investigated. Evidence was provided that those phenols which enhanced the oxidation of estradiol exerted their effect by activating peroxidase or protecting the enzyme from inactivation by the products of the reaction rather than by inhibiting the breakdown of hydrogen peroxide by catalase (EC 1.11.1.6). It has also been shown that tyrosine acted both as an activator of uterine proxidase and as a water-soluble acceptor for the metabolites of estradiol. The ability of tyrosyl peptides to form conjugates with estradiol was influenced by the other amino acids and decreased with the number of adjacent tyrosyl residues.
Cross-linking of Hemoglobin, Haptoglobin, and Hemoglobin–Haptoglobin Complex with Bifunctional Imidoesters
Authors: Lockhart W. L. , Smith David B.
Published: December 2009

Dimethyl adipimidate was used to cross-link the polypeptides within hemoglobin, haptoglobin, and hemoglobin–haptoglobin complex. Cross-linked hemoglobin retained considerable ability to bind haptoglobin, although the amounts bound were reduced and the haptoglobin reaction could be used to fractionate the modified hemoglobin. With cross-links limited to intramolecular sites, hemoglobin showed four bands on polyacrylamide gel electrophoresis in sodium dodecyl sulfate, identified, with reference to the subunit polypeptides, as monomer, dimer, trimer, and tetramer. The dimer region consisted of at least two separable species. When hemoglobin–haptoglobin complex was cross-linked, a band of hemoglobin dimer was present, which demonstrates that at least two hemoglobin subunits have a close spatial relation when bound to haptoglobin.Some comparisons with adipimidate-reacted hemoglobin were made using malonimidate and suberimidate and some marked differences were noted.
Kinetic Studies on Soluble and Insoluble Urokinases
Authors: Capet-Antonini F. C. , Tamenasse J.
Published: December 2009

A water-insoluble urokinase (ins-UK) was prepared by covalent coupling to an electrostatically neutral polyacrylamide derivative. The esteratic activity retained by the bound enzyme is about 70% of that of the soluble urokinase (UK).Comparative kinetic studies of these two forms of the enzyme were undertaken on lysine esters: N-α-acetyl-L-lysine-methyl ester (ALMe) and N-α-acetylglycyl-L-lysine methyl ester (AGLMe). It was first observed that these substrates both exhibit a marked inhibitory effect toward soluble UK, whereas this phenomenon was less manifest with the insoluble form of the enzyme. Michaelis constants and maximal velocities measured at 33 °C, for UK and ins-UK, were identical when ALMe was used, but slightly different with AGLMe. Determination of initial velocities, at a series of pH values shows only minimal differences in the behavior of the soluble enzyme with respect to that of the insoluble form. However, over a range of temperatures, differing Km values for these two enzyme forms were obtained using AGLMe as the substrate. These last results suggest possible interactions between the substrate and the insoluble carrier of the enzyme.
Isolation and Characterization of J-chain from Bovine Colostral Immunoglobulin M
Authors: Komar R. , Mukkur T. K. S.
Published: December 2009

Purified bovine colostral intact immunoglobulin M (IgM) exhibited the presence of an anodal, single, fast moving band (noncovalently bound form) when subjected to analytical polyacrylamide gel electrophoresis at an alkaline pH in urea. Reduced and alkylated or sulfitolysed bovine colostral IgM (devoid of the noncovalently bound form) also showed the presence of a similar band (covalently bound form). The molecular weight of both the covalently bound and noncovalently bound forms of the fast component was determined to be 16 500 by sodium dodecyl sulfate – polyacrylamide gel electrophoresis. In addition, the non-covalently bound form of the fast-moving component was found to be antigenically identical to the covalently bound form. The noncovalently bound form sedimented as a single peak at 1.56 S. Antiserum against the fast-moving component precipitated neither bovine colostral IgG nor μ-chains and bovine serum albumin, but precipitated native or denatured intact IgM (devoid of the non-covalently bound form) and human J-chains and vice versa, thus permitting the fast-moving components to be classified as J-chains. Radioalkylation experiments revealed the presence of 9.7 sulfhydryl groups per mole, for both the covalently and non-covalently bound forms of bovine J-chain. The stoichiometry of J-chain, determined from the densitometric tracing of the reduced and alkylated bovine colostral IgM (devoid of the noncovalently bound J-chain) in stained analytical polyacrylamide gels, revealed the presence of one J-chain per IgM molecule. On the other hand the amount of non-covalently bound form of J-chain was determined to be 1.2 per molecule of IgM.
The Transfer of Glucose to Steroids by Chimpanzee Liver Microsomes
Authors: Labow Rosalind S. , Williamson Denis G. , Layne Donald S. , Collins Delwood C.
Published: December 2009

Microsomal preparations from chimpanzee liver can transfer glucose from UDPglucose to the 17α-hydroxyl group of 17α-estradiol and of 17α-estradiol-3-glucuronide. A phenolic glucoside of estrone, but not of either 17α- or 17β-estradiol is also formed. No formation of glucosides, of p-nitrophenol, or of diethylstilbestrol was demonstrated. The specificity of glucosyl transfer in the chimpanzee is not identical to that in either the human, the rabbit, or the sheep.
Effect of Valine on the Control of Fatty Acid Synthesis in White Adipose Tissue of the Rat
Authors: Taylor Wayne M. , Halperin Mitchell L.
Published: December 2009

In adipocytes from fed rats, the rate of fatty acid synthesis in the presence of glucose and insulin was inhibited 40% by valine (5 mm). This inhibition was largely abolished by the addition to the incubation medium of the transaminase inhibitor aminooxy acetate, and of pyruvate and agents which raise the intracellular pyruvate levels such as N,N,N1,N1-tetramethyl-p-phenylenediamine.Pyruvate output into the incubation medium from fat pads obtained from fed rats and incubated with glucose and insulin was decreased significantly by the addition of valine. When adipocytes were incubated under similar conditions, the final concentration of pyruvate in the incubation medium was 42 ± 1.6 μM under control conditions and approximately one third of this value in the presence of 2.5 mM valine.Valine had no significant effect on pyruvate dehydrogenase (lipoate) (EC 1.2.4.1) activity when assayed in homogenates prepared from adipose tissue previously incubated for 60 min with the amino acid. Although the ketoacid analogue of valine, α-ketoisovaleric acid, is a competitive inhibitor of pyruvate dehydrogenase (lipoate) (Ki = 1.4 mM), this cannot solely account for the valine-induced reduced rate of lipogenesis. Rather, the mechanism involves a reduction in pyruvate concentration and thereby a diminished flow through pyruvate dehydrogenase (lipoate). Details of the possible mechanism are discussed.
A Model for The Mode of Action of Cytochalasin B Inhibition of D-Glucose Transport in the Human Erythrocyte
Authors: Taylor N. F. , Gagneja G. L.
Published: December 2009

By an optical method, cytochalasin B is shown to be a competitive inhibitor of D-glucose transport across the human erythrocyte membrane with Ki of 1.2 × 10−7 M. A Drieding molecular model of cytochalasin B reveals an almost identical spatial distribution of four oxygen atoms to those found in the C1-conformation of β-D-glucopyranose and implicated in hydrogen bonding to the carrier protein associated with D-glucose transport. The stereochemistry of this transport model is discussed.On the basis of the interoxygen distances found in cytochalasin B, hydrocortisone, prednisolone, corticosterone, and phenolphthalein are considered as analogues and are shown to be competitive inhibitors of D-glucose transport with Ki values of 2.2 × 10−4 M, 3.0 × 10−4 M, 4.0 × 10−4 M, and 2.5 × 10−5 M, respectively. These results are considered to be consistent with the proposed mode of action of cytochalasin B and also provide further support for the model of D-glucose stereospecifically hydrogen-bonded to a carrier protein.
Porcine Chymotrypsin A-π, a More Acidic Chymotrypsin
Authors: Médicis Eveline de , Bergeron Louise
Published: December 2009

A kinetic study of porcine chymotrypsin A-π revealed two characteristic properties of this type of chymotrypsin:1. Porcine chymotrypsin A-π, like bovine chymotrypsin B-π, does not bind proflavin. which is a competitive inhibitor of bovine trypsin and chymotrypsin A-α.2. The pH profiles of the steady-state parameters show the two usual important pK's. The basic one, pK2 = 9.6, affects both Km and kcat/Km and probably controls the binding conformation of chymotrypsin. The acidic one, pK1 = 5.7, affects kcat and kcat/Km and plays a role in the catalytic process. The value of pK1 is unusually low.