bluuebunny/crossref_metadata_2025 huggingface dataset:
Effect of temperature and current manipulation on smolting in Chinook salmon (Oncorhynchus tshawytscha): the relationship between migratory behaviour and physiological development
Authors: Sykes Gregory E. , Shrimpton J. Mark
Published: December 2009

We investigated the role of temperature and current on migration pattern and physiological change in Chinook salmon ( Oncorhynchus tshawytscha ) smolts. Chinook parr were implanted with passive integrated transponder tags and placed in circular tanks with directional current. Paired antennas in each tank allowed for the monitoring of movements with or against the current. Two temperature regimes, naturally increasing (warm) and constant (cool), were combined with high current (>0.5 m·s–1) and low current (<0.1 m·s–1) to create four experimental treatments. At 2-week intervals, fish were sampled for gill Na+,K+-ATPase activity and plasma cortisol concentration. Increasing temperature resulted in an earlier peak in movement than colder groups and showed a distinct peak in gill Na+,K+-ATPase activity. Current did not influence physiological parameters associated with smolting and was not found to initiate movement; movement was related to accumulated thermal units (ATU). The presence of a strong, directional current, however, resulted in a period of more defined movement, suggesting a possible influence of current once migration is underway. A combination of either photoperiod or ATU with gill Na+,K+-ATPase was most strongly linked to movement. ATU was also found to be more strongly correlated with the smolting process than daily mean temperature.
Quantifying thermal effects on contemporary growth variability to predict responses to climate change in northern rock sole (Lepidopsetta polyxystra)
Authors: Hurst Thomas P. , Abookire Alisa A. , Knoth Brian
Published: December 2009

Understanding the causes of contemporary variation in growth rates can offer insights into the likely consequences of climate change for growth and recruitment of coastal marine fishes. We examined the growth dynamics of age-0 northern rock sole ( Lepidopsetta polyxystra ) over four years in three nurseries at Kodiak Island, Alaska, USA. Following the settlement period, fish were sampled monthly (July–October) with a 3 m beam trawl at fixed-position transects. Postsettlement sizes were positively related to temperatures during the spawning and larval periods, suggesting environmental control of spawning or settlement timing. Summer growth on the nursery grounds varied significantly among sites and years (mean size 32.8–63.1 mm in mid-September), with the Holiday Beach site consistently supporting the fastest growth rates. Contrary to expectations of density dependence and thermal regulation, nursery ground growth rates were not significantly correlated with fish density or water temperatures. The minor contribution of thermal variation to growth rates appears related to the conservative growth strategy and low thermal sensitivity of northern rock sole. These results suggest that climate changes influencing spawning time and larval growth may have larger impacts on first-year growth and recruitment of this species than temperature effects on the growth of nursery-resident juveniles.
SYNTHESIS OF SHIKIMIC ACID-C14IN PINUS RESINOSA, AIT. FROM SOME C14-LABELLED COMPOUNDS
Authors: Yoshida Seiichi , Towers G. H. N.
Published: December 2009

The shikimic acid (SA) content of Pinus resinosa twigs was found to range from 0.2–2.12 g fresh weight, depending on the season of the year. Of the radioactive compounds administered to cut shoots, pyruvic acid gave SA of the highest specific activity. Sugars were next most effective, with glucose and ribose being better than sucrose and fructose. Carbon from several organic acids was not readily incorporated into SA. Pyruvate-3-C14gave SA of higher specific activity than did pyruvate-1-C14or -2-C14. When pyruvate-1-C14was used, 40% of the activity in shikimic acid was in the carboxyl carbon, whereas with pyruvate-2-C14or -3-C14only 3–10% of it was in the carboxyl carbon. The results with pyruvate can best be explained in terms of the "dicarboxylate shuttle". When either erythrose or 5-dehydroshikimic acid was administered together with pyruvate-C14, a decrease in specific activity of SA was observed. These results are consistent with the view that in Pinus, SA is synthesized from phosphoenolpyruvate and erythrose-4-phosphate through dehydroshikimic acid.
ACTIVE TRANSPORT OF ASCORBIC ACID IN ADRENAL CORTEX AND BRAIN CORTEX IN VITRO AND THE EFFECTS OF ACTH AND STEROIDS
Authors: Sharma Shail K. , Johnstone R. M. , Quastel J. H.
Published: December 2009

Uptake of ascorbic acid-1-C14in brain cortex and adrenal cortex slices is an energy-dependent process. Concentration ratios (i.e. ratios of tissue ascorbic acid-1-C14to medium ascorbic acid-1-C14) greater than 4 have been obtained with both tissues in vitro. Ouabain as well as 2, 4-dinitrophenol suppresses ascorbic acid uptake into brain cortex slices.ACTH inhibits the uptake of ascorbic acid-1-C14in adrenal cortex, but not in brain cortex slices. The presence of glucose is necessary for the inhibition. Several cortical steroids, as well as adenosine-3′,5′-monophosphate, at small concentrations inhibit the uptake. The results are consistent with the interpretation that ACTH inhibits the uptake of ascorbic acid in the adrenal cortex through the steroids produced in its presence.
STUDIES OF LIGNIN BIOSYNTHESIS USING ISOTOPIC CARBON: XIII. THE PHENYLPROPANOID SYSTEM IN LIGNIFICATION
Authors: Higuchi Takayoshi , Brown Stewart A.
Published: December 2009

Techniques of isotope competition and trapping were used to study the phenylpropanoid biosynthetic pathway in lignifying wheat plants. The results in general confirm earlier findings that phenyllactic acid (PLA), p-hydroxyphenyllactic acid (HPLA), phenylpyruvic, cinnamic, caffeic, ferulic, and sinapic acids can participate in lignification. L-Phenylalanine and L-tyrosine were converted to PLA and HPLA, respectively, but there was much less conversion of cinnamic acid to PLA, or p-hydroxycinnamic acid to HPLA. A pathway from phenylalanine to cinnamic acid via PLA, and an analogous pathway involving tyrosine thus remain as possible alternatives to the established routes involving deamination of these amino acids by phenylalanine deaminase or tyrase. Feeding of non-radioactive coniferyl alcohol with ferulic acid-C14results in the formation of both coniferyl- and sinapyl-type lignin residues having lower specific radioactivities than were obtained after the feeding of ferulic acid-C14alone. After a 5-hour metabolic period in the presence of ferulic acid-C14, both coniferyl aldehyde and coniferyl alcohol became labelled, and the radioactivity of the aldehyde was much higher than that of the alcohol. There was no evidence of coniferin formation. These findings indicate that coniferyl alcohol is formed from ferulic acid through coniferyl aldehyde, and that coniferin is probably unnecessary for lignification, at least in species other than conifers.
IMPROVED PROCEDURES FOR PREPARATION AND CHARACTERIZATION OF MYROTHECIUM CELLULASE: PART 4. CHARACTERIZATION OF ACTIVITY TOWARD β-METHYL GLYCOSIDES OF 1 → 4-β-D-OLIGOGLUCOSIDES
Authors: Hanstein Else G. , Whitaker D. R.
Published: December 2009

Two methods for the preparation of β-methyl glycosides of 1 → 4-β-D-oligoglucosides are described. In method I, the parent oligoglucoside is acetylated, acetobrominated, methylated, fractionated on Magnesol, and deacetylated. In method II, an acetolysate of cellulose is acetobrominated, methylated, deacetylated, and fractionated on Darco G60 charcoal.The parameters Kmand k2for the hydrolysis of β-methyl cellotetraoside by purified Myrothecium cellulase at pH 5.0 and 28.6° were estimated to be 4.1 × 10−4 M and 570 minute−1respectively. Kmdecreases and k2increases as the D.P. of the substrate is increased from three to five. The initial hydrolysis products of β-methyl cellotetraoside and β-methyl cellopentaoside indicate that the interior but not the terminal linkages are hydrolyzed by the enzyme.
MECHANISM OF ACTIVATION OF HOG PANCREATIC LIPASE BY SODIUM TAUROCHOLATE
Authors: Fritz Paul J. , Melius Paul
Published: December 2009

A new preparation of hog pancreatic lipase is described which is evidently the most stable one of its kind. From the composition of the reaction mixtures after enzymatic hydrolysis with and without sodium taurocholate, it appears that the hydrolysis of triglyceride to diglyceride is facilitated and the hydrolysis of diglyceride to monoglyceride is depressed in the presence of taurocholate. The differences in the curves showing the rates of hydrolysis of triolein, monoolein, tributyrin, and monobutyrin in the presence and absence of taurocholate also indicate that the taurocholate acts to split the diglyceride–enzyme complex and thus increases the action of the enzyme on the triglyceride ester. The hydrolysis of the triglyceride is a fast reaction whereas the di- and mono-glycerides are hydrolyzed at much slower rates. The activation energy for the lipolysis of the triolein and tributyrin has been calculated in the presence and absence of taurocholate. This was possible because in spite of earlier reports that the lipolysis of triolein was not temperature dependent between 10 and 40 degrees, we found these reactions to be temperature dependent. The lesser activating effect at the highest taurocholate concentrations indicates this is not a simple emulsifying effect.
AUTOTROPHIC ENZYME SYSTEMS: I. ELECTRON TRANSPORT SYSTEMS CONCERNED WITH HYDROXYLAMINE OXIDATION IN NITROSOMONAS
Authors: Aleem M. I. H. , Lees H.
Published: December 2009

Intact cells or cell-free extracts of Nitrosomonas catalyze the rapid and stoichiometric conversion of nitrohydroxylamine to nitrite at rates comparable to the oxidation of ammonium ion or hydroxylamine to nitrite. Cell-free extracts possess a powerful hydroxylamine – cytochrome c reductase activating hydroxylamine to donate electrons to the cytochrome systems comprising b, c, and a type components. The partially purified enzyme complex is sensitive to low concentrations of cyanide and inhibitors of the flavoproteins. The possible mechanism of the formation and oxidation of the new intermediate "nitrohydroxylamine" in the reaction sequence is discussed.
DEAMINATION OF ADENINE AND RELATED COMPOUNDS AND FORMATION OF DEOXYADENOSINE AND DEOXYINOSINE BY LINGCOD MUSCLE ENZYMES
Authors: Tarr H. L. A. , Comer A. G.
Published: December 2009

Adenylic acid deaminase was destroyed by heating crude extracts of lingcod muscle to 58 °C, while deamination of adenosine and deoxyadenosine was unaffected. Deoxyadenosine was deaminated about 2300 times as fast as adenine by the enzyme extract. The specific activity of the purine nucleoside phosphorylase with adenine and deoxyribose-1-phosphate as substrates was similar to that of the deoxyadenosine deaminase of the enzyme preparations. It can thus be stated with certainty that adenine is a substrate for the nucleoside phosphorylase, that deoxyadenosine thus formed is promptly deaminated to deoxyinosine, and that deamination of adenine to hypoxanthine plays a negligible role in formation of deoxyinosine. The presence of small amounts of deoxyadenosine in the above reaction was verified by radioactive adenine and isolation of deoxyadenosine of constant specific activity by addition of deoxyadenosine carrier.
SYNTHESIS OF PHOSPHOLIPID METABOLITES:L-α-GLYCERYLPHOSPHORYL-2-AMINO-2-METHYLPROPANOL
Authors: Baer Erich , Rao G. Venkat
Published: December 2009

A constituent of and possible intermediate in the metabolism of phosphatidyl-2-amino-2-methylpropanols has been synthesized. The fatty-acid-free moiety, i.e. L-α-glycerylphosphoryl-2-amino-2-methylpropanol, was obtained by phosphorylating D-acetone glycerol with phosphorus oxychloride and quinoline, esterifying the acetone L-α-glycerylphosphoric acid dichloride with N-carbobenzoxy-2-amino-2-methylpropanol in the presence of pyridine, removing the acetone group of acetone L-α-glycerylphosphoryl-N-carbobenzoxy-2-amino-2-methylpropanol by mild acid hydrolysis, and freeing the barium salt of L-α-glycerylphosphoryl-N-carbobenzoxy-2-amino-2-methylpropanol from its protective carbobenzoxy group by catalytic hydrogenolysis.
ALDOLASE ACTIVITY IN CONFUSED FLOUR BEETLE, TRIBOLIUM CONFUSUM DUVAL (TENEBRIONIDAE, COLEOPTERA)
Authors: Chaudhary K. D. , Srivastava U. , Lemonde A.
Published: December 2009

Certain biochemical characteristics of aldolase in Tribolium confusum homogenate have been determined. The Michaelis constant (Km) has been calculated to be 1.2 × 10−3 M. The effect of various inhibitors suggests that T. confusum aldolase resembles muscle aldolase.Intracellular localization of the enzymatic activity shows that aldolase is contained mainly in the soluble fraction and to a less extent, in an unbound form, in nuclei.Aldolase activity has been demonstrated in relation to various stages in the life cycle of this insect. Such a study reveals that the Embden-Meyerhof glycolytic pathway is not a prominent energy-yielding pathway in T. confusum.
SYNTHESIS OF NUCLEOSIDE DIPHOSPHATES AND TRIPHOSPHATES BY A SALMON MILT ENZYME PREPARATION
Authors: Tarr H. L. A. , Roy Joan
Published: December 2009

A cell-free enzyme preparation capable of forming nucleoside (or deoxynucleoside) di- and tri-phosphates from the corresponding nucleoside monophosphates in the presence of adenosine triphosphate (ATP) was prepared from immature salmon milts. The preparation was not free from phosphatase activities. The corresponding nucleoside (or deoxynucleoside) di- and tri-phosphates were readily formed from cytidine monophosphate, adenosine monophosphate, uridine monophosphate, and deoxyuridine monophosphate; guanosine monophosphate, deoxycytidine monophosphate, inosine monophosphate, and deoxyribosylthymine monophosphate were poorer substrates. The reaction required ATP. Guanosine triphosphate, uridine triphosphate, cytidine triphosphate, deoxyribosylthymine triphosphate, and inosine triphosphate could replace ATP, though they were in general less effective. With cytidine monophosphate as substrate, the formation of cytidine triphosphate was accelerated by Mg++, Ca++, Mn++, Co++, Ni++, and CN−, and was inhibited by Cu++, Ag+, Zn++, Hg++, F−, iodoacetate, p-hydroxymercuribenzoate, and ethylenediaminetetraacetate. Neither glutathione nor 2-mercaptoethanol in concentrations of between 1 × 10−3 and 1 × 10−1 M protected the enzyme. The optimum pH was 7.5–8.0. The enzyme was very unstable, losing two-thirds of its activity in 1 hour at 30 °C. Under optimum conditions with cytidine monophosphate as substrate, 8.5 μmoles of cytidine triphosphate were formed per hour per milligram of protein nitrogen at 25 °C.
Characterization of Two Polygalacturonases Produced by Verticillium albo-atrum
Authors: Mussell H. W. , Strouse Blanche
Published: December 2009

Two polygalacturonases (poly-α-1,4-galacturonide glycanohydrolase, EC 3.2.1.15) from the culture fluids of Verticillium albo-atrum were purified and characterized. Gel permeation and exchange chromatography followed by double isoelectric focusing have been used to separate and purify an exopolygalacturonase and an endopolygalacturonase. The exopolygalacturonase had a molecular weight of 44 000, pI of 6.5, and pH optima of 4.5 for pectin and 5.25 for sodium polypectate. The endopolygalacturonase had a molecular weight of 34 500, pI of 9.7, and pH optima of 5.0 for pectin and 6.0 for sodium polypectate. Both purified enzymes were electrophoretically homogeneous on polyacrylamide gels and exceptionally stable when stored in solution at 4°. The endo-enzyme was 20 times as effective in macerating plant tissue as the exo-enzyme.
Purification of Fully Active Human Encephalitogenic Basic Protein on Sulfoethyl-Sephadex
Authors: Barton M. A. , McPherson T. A. , Martin J. K.
Published: December 2009

Human encephalitogenic basic protein (HEP) was obtained in high yield after chromatography on sulfoethyl-Sephadex C-50 (SE-Sephadex). The protein was shown to be homogeneous and was identical in physical properties with samples of human encephalitogenic protein obtained by other methods. It was also shown to induce typical clinical and histological experimental allergic encephalomyelitis (E.A.E.) in adult guinea pigs injected with 50 μg HEP in Freund's complete adjuvant (FCA). Gel-filtration radioimmunoassay was used to study immunogenicity, antigenicity, and antigenic cross-reactivity of HEP; seven of 11 guinea pigs injected with HEP in FCA had detectable serum antibody to 125I-HEP, indicating that the SE-Sephadex preparation was fully immunogenic and antigenic; HEP and another preparation of human encephalitogenic basic protein prepared by a different method were equally capable of inhibiting the binding between 125I-HEP and anti-HEP serum indicating that the two samples tested were antigenically identical. We believe that the new technique represents a significant technological advance because it provides a most effective and rapid method for the isolation of fully active encephalitogenic basic protein.
Endogenous Alanine, Glutamate, Aspartate, and Glutamine in the Perfused Guinea-Pig Heart: Effect of Substrates and Cardioactive Agents
Authors: Gailis L. , Benmouyal E.
Published: December 2009

The isolated, nonworking guinea-pig heart was perfused with 14C-labelled glucose, pyruvate, and acetate. Labelled and total alanine, glutamate, aspartate, and glutamine were measured. The alanine content of the heart varied directly with the medium pyruvate concentration. The sum of the concentrations of glutamate and aspartate varied inversely with the alanine concentration. The sum of alanine, glutamate, aspartate, and glutamine in the heart was constant under most perfusion conditions, but loss into the medium increased in the presence of ouabain, low Ca2+ concentration, or malonate. The sum of the total amino acids from the heart and medium remained constant under all conditions except in the presence of malonate, when it was decreased.Labelling of amino acids from 14C-labelled substrates indicated that alanine, glutamate, and aspartate exchanged readily with their α-oxo acids. However, only 30% of the glutamine exchanged with glutamate during 1 h; this proportion varied little with perfusion conditions or the metabolic flux. The results indicate that in a closed system, most of the changes in the concentrations of amino acids are brought about by transamination.
Succinyl Coenzyme A Synthetase of Escherichia coli: Initial Rate Kinetics of Succinyl-CoA Cleavage and Isotope Exchange Studies
Authors: Moffet Frank J. , Bridger W. A.
Published: December 2009

Initial rate kinetic studies of succinyl coenzyme A synthetase of E. coli in the direction of succinyl-CoA cleavage are consistent with the operation of a partially random sequential kinetic mechanism with initial binding of ADP followed by random association of succinyl-CoA and Pi. The mechanism is analogous to that proposed previously for the succinyl-CoA formation reaction, and thus the kinetic mechanism of the overall reversible succinyl-CoA synthetase reaction appears to be symmetrical.Studies of the kinetics of [Formula: see text] isotope exchange at equilibrium show that this partially random sequential kinetic mechanism is not an exclusive pathway. [Formula: see text] isotope exchange rates did not show complete substrate inhibition when CoA or succinate was varied in constant ratio with Pi. However, when CoA or succinate was varied in constant ratio with succinyl-CoA, nearly complete substrate inhibition was observed. These results can be interpreted in terms of a wide variety of minor pathways of substrate binding and product release available to the enzyme under various conditions.
Temperature Sensitivity of Myosin and Actomyosin
Authors: Jacobson A. L. , Henderson J.
Published: December 2009

The thermal denaturation of myosin and actomyosin was studied by active site analysis (enzymatic activity) and measurements were related to overall conformational changes (viscosity) over the temperature range 19–65 °C. The role of sulfhydryl (SH) groups on the temperature-induced denaturation of actomyosin was investigated. The temperature of maximum change in the overall conformation (the melting temperature, TM) was unaffected by the binding of F-actin to myosin. For both myosin and actomyosin the TM was 43 ± 2 °C. However, the range of temperature over which large conformational changes were observed was affected by the binding of F-actin to myosin. For myosin and dissociated actomyosin, changes were observed between 37 and 50 °C, while for actomyosin large changes were observed between 20 and 50 °C. With actomyosin there was an irreversible increase in titratable sulfhydryl groups from 19 to 60 °C. Temperature effects on the calcium-activated ATPase were studied. The temperature of maximum enzymatic activity for actomyosin was 45 ± 2 °C, which corresponds to the TM and the temperature at which increases in SH content were apparent. However, for myosin the temperature of maximum enzymatic activity was 33 °C, considerably below the TM. Overall conformational changes were reversible below the TM, while changes in enzymatic activity were reversible below the temperature of maximum enzymatic activity. Actin offers considerable protection to the temperature inactivation of the active site of myosin even though the F-actin–myosin complex is very highly dissociated in the presence of ATP. However, there is no significant stabilization of myosin by F-actin in terms of the temperature sensitivity of the overall conformation.
Amount of 14C Assimilated in Different Organs and Tissues of the Rat After a Single Injection of [U-14C]glucose
Authors: Vrba R. , Winter Anna
Published: December 2009

Labelling of proteins and lipids of 20 organs and tissues of the rat was studied 4 h after subcutaneous injection of D(+)-[U-14C]glucose. Gross differences between the rate of labelling of acid-insoluble constituents of various organs were noted; specific radioactivities of protein fractions obtained from the pancreas, plasma, ileojejunal segment of the alimentary tract, thymus, and spleen were equal to or higher than those of the brain protein fraction. This indicates that, apart from the brain, the high rates of assimilation of glucose carbon occur at anatomical sites of release of proteins, of rapid cell proliferation, or of immunological responses. In comparison with the above-mentioned organs, the rate of incorporation of glucose carbon into protein fractions of the heart, skeletal muscles, and formed elements of blood was low. In all organs and tissues investigated significantly more glucose carbon was incorporated into protein fractions than into lipid fractions, with the exception of adipose tissue, where glucose carbon was incorporated mainly into the lipid fraction.
Extramitochondrial Location of the Minor Alanine-Specific tRNA Stimulatory to Hemoglobin Formation in the Chick Blood Islands
Authors: Wainwright S. D. , Wainwright Lillian K.
Published: December 2009

Preparations of tRNA isolated from chicken liver mitochondria did not stimulate formation of hemoglobin by chick blastodiscs explanted on medium containing actinomycin D. Profiles of liver mitochondrial alanyl-tRNA on columns of benzoylated diethylaminoethylcellulose were almost indistinguishable from those of chick embryo alanyl-tRNA, but did appear to contain a smaller proportion of the minor species believed to stimulate hemoglobin formation.
Studies on the Alkaline Denaturation of Glucagon
Authors: Epand Richard M. , Epand Raquel F.
Published: December 2009

In solutions at high pH, amino acid residues of glucagon undergo racemization without cleavage of the polypeptide backbone. This racemization process is accompanied by an unfolding of the molecule with the loss of both its antigenicity and its ability to stimulate adenyl cyclase from rat liver.
Binding of Actinomycin Chromophore Analogues to DNA
Authors: Chibber B. A. K. , Balasubramanian A. , Viswanatha T.
Published: December 2009

Five chromophore analogues of actinomycin, viz. phenoxazone (1), aminophenoxazone (2), N-acetylaminophenoxazone (3), N-methylaminophenoxazone (4), and actinocin (5), were synthesized. Of these compounds, only 2 and 4 were found to interact with DNA. The binding constants for these compounds, obtained by spectrophotometric titration, were of the same order of magnitude as that obtained with actinomycin-DNA interaction. The major difference between the binding of actinomycin and of the analogues to DNA was the lability of the complex of the latter compounds in the presence of Mg2+ ions. These observations have been interpreted to indicate intercalation of the chromophore analogues between successive base pairs of DNA. The spectra of actinocin were unaffected by the addition of DNA. The spectral changes indicative of an interaction between actinocin and DNA observed at low concentration of Mg2+ ions were abolished at high concentrations of these ions. The implications of these observations to the mechanism of actinomycin binding to DNA are discussed.
The Half-Molecule of Haptoglobin: Studies on the Product Obtained by the Selective Cleavage of a Haptoglobin Disulfide
Authors: Malchy B. , Rorstad O. , Dixon G. H.
Published: December 2009

A disulfide of haptoglobin 1-1 has been selectively cleaved using sodium sulfite and p-chloro-mercurisulfonate. The reaction product (half-haptoglobin) could be separated from haptoglobin by gel electrophoresis in urea or sodium dodecyl sulfate, and its molecular weight was one-half that of native haptoglobin 1-1. When the sulfite cleavage reaction was performed on haptoglobin 2-2 the haptoglobin polymers were broken down. Isolation of the S-sulfocysteine-containing peptide revealed that the disulfide bond broken in the formation of half-haptoglobin was the 21α-21α disulfide. This conclusion was confirmed by analysis of the cyanogen bromide fragments from half-haptoglobin and by a performic acid diagonal analysis of peptic peptides obtained from half-haptoglobin.
Fluorescence Quenching, a Tool for Probing Conformational Changes in Glycogen Phosphorylase
Authors: Honikel K. O. , Madsen N. B.
Published: December 2009

This study shows that conformational changes in glycogen phosphorylase are accompanied by changes in the accessibility of tryptophan residues and of the coenzyme, pyridoxal phosphate, to the surrounding aqueous medium. The accessibility was estimated by determining the extent to which iodide can quench the fluorescence emission of these moieties by colliding with them, since iodide cannot collide with a buried chromophore and hence cannot quench its fluorescence. Rabbit muscle phosphorylase b, its apoform, and phosphorylase a exhibit differences in the number of exposed tryptophans, while the phosphorylase b forms from rabbit skeletal muscle and pig heart also show differences.Differences are also observed in the accessibility of the coenzyme in different forms of the enzyme. The quenching rate constant, a measure of accessibility, differs for phosphorylases a and b, and this constant is affected differently by ligand binding to the two forms. While the allosteric inhibitors, ATP and glucose 6-phosphate, render the pyridoxal phosphate moiety of phosphorylase b more accessible, the activator, AMP, and substrate, glucose 1-phosphate, together cause it to be totally inaccessible to fluorescence quenching by iodide. AMP and glucose 1-phosphate appear to mediate a conformational change which buries the coenzyme. While pyridoxal phosphate is necessary for catalytic activity, one may conclude from these experiments that its ring structure is unlikely to participate directly in the catalytic mechanism.
The Partial Purification and Properties of Uridine Kinase from Ehrlich Ascites Tumor Cells
Authors: Krystal Gerald , Scholefield Peter G.
Published: December 2009

In the partial purification of uridine kinase (ATP:uridine 5′-phosphotransferase, EC 2.7.1.48) from Ehrlich ascites tumor cells, two active fractions were obtained by chromatography on Sepharose 6B. Their molecular weights, as determined by a gel filtration and sucrose density centrifugation, were approximately 120 000 and 30 000. The larger molecular weight species was less stable on incubation at 60° and slightly less sensitive to CTP inhibition than the lower molecular weight fraction. Both fractions were stabilized against heat inactivation by ATP and CTP.The two fractions displayed similar apparent Km values for uridine and ATP, similar specificities for substrates and inhibitors, and similar divalent cation requirements.In the partial purification of uridine kinase from mouse intestine, only the larger molecular weight fraction was obtained by chromatography on Sepharose 6B.In addition, some preliminary evidence for a monomer–polymer relationship between the two molecular weight species is presented.
Perfusion of the Boar Testis with Androstenediol Sulfate in vivo and Studies on the in vitro Metabolism of Androgens by Porcine Blood Cells
Authors: Kraemer M. A. , Raeside J. I.
Published: December 2009

An attempt was made to see if androstenediol sulfate, a normal secretory product of the boar testis, is important in testicular steroidogenesis in this species when perfused in vivo through the testis. Approximately 5 μCi of 7α-3H-androstenediol sulfate were administered via the spermatic artery to four, mature, anesthetized boars. Almost all of the radioactivity extracted from the spermatic vein blood and from the testis was associated with the unaltered substrate. Less than 0.2% of the substrate was isolated as 3H-dehydroepiandrosterone sulfate from the testicular extracts of one boar. An unsuccessful effort was made to characterize a similar amount of a metabolite present in extracts from spermatic vein blood of the same animal. The total amount of radioactivity which was associated with the free steroids represented about 0.2% of the administered dose of 3H-androstenediol sulfate given to each boar.17β-Hydroxysteroid dehydrogenase activity was demonstrated in a series of incubations of steroids with blood cells of the boar. Unconjugated Δ5-steroids were converted to a greater extent than were the Δ4-steroids (3.33 and 3.91% for DHA and androstenediol, respectively; 1.23 and 0.92% for androstenedione and testosterone, respectively). Androstenediol sulfate was converted to dehydroepiandrosterone sulfate (0.42%) but the reverse reaction was not observed. These findings were considered in relation to the in vivo studies. It was concluded that androstenediol sulfate in arterial blood entering the boar testis was not metabolized appreciably.
Lactobacillus Plantarum Exoribonuclease: The Effect of Partial Tryptic Digestion on Enzyme Kinetics and the Mode of Degradation
Authors: Gardonio E. C. , Logan D. M.
Published: December 2009

The exoribonuclease from Lactobacillus plantarum was subjected to partial tryptic digestion to test the effect of primary structure modification on the enzymes processive mode of degradation. Tryptic digestion leads to large decreases in both Km and Vmax but these kinetic changes can be reduced if the enzyme is incubated with a suitable substrate such as poly A prior to trypsin treatment. In addition trypsin treatment leads to an enzyme with modified ion requirements and anomalous substrate dependence. Substrate dependence curves show both hyperbolic and sigmoidal sections which may be related to modification of nonbinding and binding positions, respectively, in the enzyme. Hill plots of the data reveal multiple binding sites for substrates such as poly A. When the enzyme is treated until less than 20–30% activity remains, the previous strict processivity of action is lost and a short oligonucleotide is produced which is then degraded processively.
Characterization of Canine Hepatic and Renal Esterases
Authors: Ecobichon D. J.
Published: December 2009

The esterases of canine liver and kidney were separated electrophoretically into nine bands with identical migration patterns in both tissues. An additional pair of rapidly migrating anodic bands were observed in hepatic extracts. Based on substrate specificity, the predominant tissue esterases were identified as nonspecific carboxylesterases (aliesterases). No cholinesterase activity was detected in the tissue extracts. Kinetic characteristics determined for the hepatic and renal esterases included (1) optimal pH; (2) Km values for esters of α-naphthyl and p-nitrophenol; (3) average rates of hydrolysis of α-naphthyl acetate and p-nitrophenyl acetate by the tissue extracts. Inhibition studies revealed the presence of two types of esterase activity in each tissue: one type being sensitive to organophosphorus esters, the second being resistant. A study of preferential substrate hydrolysis in the presence of known characteristic activators and inhibitors of esterases revealed approximately 5% and 20% arylesterase activity in liver and kidney, respectively. The presence of arylesterase activity in these tissues was confirmed by the hydrolysis of paraoxon (E600).
Transcription of Satellite DNA in Mouse L-cells
Authors: Cohen A. K. , Huh T. Y. , Helleiner C. W.
Published: December 2009

Mouse L-cells contain RNA capable of hybridizing with mouse satellite DNA on nitrocellulose membranes and of competing for hybridizing sites on satellite DNA with transcript of satellite DNA prepared in vitro. RNA prepared from either whole cells or from the cytoplasm includes a fraction hybridizing with satellite DNA. That from whole cells has a half-life of 17 min, whereas that from cytoplasm is stable.
Plasma Cholesterol Esterification in Normal and Hyperlipoproteinemic Subjects
Authors: Fabien Hélène D. , Davignon Jean , Marcel Yves L.
Published: December 2009

Plasma lipid values and the initial rates of plasma cholesterol esterification in vitro have been studied in seven normal subjects and in 34 patients with primary hyperlipoproteinemia of the different types.Compared with that of normal subjects, the mean rate of cholesterol esterification (nanomoles cholesterol esterified per hour per milliliter of plasma) is significantly increased in type II (p < 0.05), whereas the mean increases noted in types III, IV, and V are not significant. The mean relative rate (percentage cholesterol esterified per hour) is decreased in type III (p < 0.05).The correlations between plasma lipid values and cholesterol esterification have been calculated in each type as well as in normal and hyperlipoproteinemic subjects taken as a group (total group). In the total group, plasma cholesterol esterification (nanomoles per hour per milliliter) is weakly correlated with unesterified cholesterol (r = 0.384), with triglycerides (r = 0.416), and with the ratio of triglycerides to total cholesterol (r = 0.439). Only in the group of normal subjects is the rate of cholesterol esterification strongly correlated with triglycerides (r = 0.896) and with the ratio of triglycerides to total cholesterol (r = 0.885). Finally, there is a strong negative correlation (r = −0.792, p < 0.001) between the rate of cholesterol esterification expressed in the percentage of cholesterol esterified per hour and the ponderal index in the total group. This negative correlation indicates a positive correlation between cholesterol esterification and obesity. In all subjects, we have also found that negative or positive caloric balances caused the rates of cholesterol esterification to be, respectively, decreased or increased as compared with the mean of the group considered.
Oxidation of Sulfur Compounds and Coupled Phosphorylation in the Chemoautotroph Thiobacillus neapolitanus
Authors: Saxena J. , Aleem M. I. H.
Published: December 2009

The intact cells from T. neapolitanus oxidized thiosulfate completely to sulfate with the consumption of 2 mol O2/mol thiosulfate. The oxidation of thiosulfate by cell-free extracts was incomplete and resulted in the accumulation of tetrathionate. Aerobic treatment of the intact cells with thiosulfate caused a rapid increase in the cellular ATP content which was unaffected by the flavoprotein inhibitors and antimycin A but was strongly inhibited by uncouplers of oxidative phosphorylation. The oxidation of sulfide by intact cells and cell-free extracts appears to proceed via two stages: (1) a cellular-membrane-associated initial and rapid oxidation stage which was dependent upon sulfide concentration, and (2) a slower oxidation reaction stage which was catalyzed by the soluble cell-free fractions. The oxidation of sulfide and 2-mercaptoethanol by intact cells or cell-free extracts was coupled to ATP generation and the process was inhibited by the respiratory chain inhibitors such as amytal, antimycin A, and cyanide, as well as by pentachlorophenol and carbonyl cyanide m-chlorophenyl hydrazone. The oxidation of sulfide and 2-mercaptoefhanol by cell-free extracts yielded P/O ratios of 0.3 and 0.85, respectively. The intact cells (in the absence of added electron donors) were able to exhibit an acid-induced ATP formation from the phosphorylation of endogenous ADP which could be inhibited effectively by uncouplers of oxidative phorylation but was unaffected by respiratory chain inhibitors.
The Denatured States of Lysozyme
Authors: Kugimiya M. , Bigelow C. C.
Published: December 2009

The denaturation of hen egg-white lysozyme has been studied under a variety of denaturing conditions by difference spectroscopy, polarimetry, and viscometry. It has been found that the lysozyme molecule can take up four different denatured conformations, depending on the denaturant used. Urea and guanidinium chloride produce the most completely denatured state, IV, and, listed in the order of increasing residual structure, lithium chloride gives state III, temperature state II, and lithium perchlorate state I. Differences in the spectra at 300 nm have been a valuable aid in relating the different denatured states to each other.
Wheat-Embryo Ribonucleates. I. Subcellular Localization of a Satellite Polyribonucleotide
Authors: Azad A. A. , Lane B. G.
Published: December 2009

(1) When wheat embryos are extracted with aqueous phenol and the aqueous phase is made 2.5 M with respect to NaCl at 0 °C, there is selective precipitation of about 80% of the total RNA. The 18 S and 26 S RNA species from the wheat-embryo ribosomes comprise a preponderant mass fraction (ca. 80%) of this NaCl-insoluble RNA (iRNA). A small amount of a rapidly migrating electrophoretic component (iRMEC) can be released by aqueous denaturation of wheat-embryo NaCl-insoluble RNA and because it is specifically complexed with 26 S RNA, the iRMEC component has been termed a "satellite" of 26 S RNA.(2) The wheat-embryo satellite RNA has been shown to be present in the microsomal fraction recovered from cell-free homogenates of wheat embryos.(3) The wheat-embryo satellite RNA has been shown to be differentially localized in the large subunit of wheat-embryo ribosomes where it presumably exists as part of the same intermolecular 26 S RNA complex that can be isolated by directly extracting the whole embryos with aqueous phenol.(4) During preparation of the ribosomal subunits, there is substantial degradation of the component ribonucleates and the nature of this degradation is the subject of a brief discussion.
Circular Dichroism Analysis of Native and Reaggregated Mycoplasma Membranes
Authors: Rottem Shlomo , Hayflick Leonard
Published: December 2009

Circular dichroism analyses of Acholeplasma laidlawii membranes solubilized by sodium dodecyl sulfate showed a typical α-helix spectrum. The estimated α-helix content was of the order of 35% calculated from the ellipticity at 208 nm of membranes solubilized by 20 mM SDS. Reaggregation of the solubilized membrane material to a membrane-like structure resulted in a distorted spectrum with low amplitude and red-shifted extrema like that of the native membranes. Throughout the growth cycle the circular dichroism spectra of membrane proteins remained the same despite the marked differences in membrane densities. The optical activity of the membranes was not affected by changing the lipid composition or extraction of over 90% of the lipids, although the latter resulted in marked destabilization of the proteins.
Fractionation of Chromatin Components
Authors: Monahan John J. , Hall Ross H.
Published: December 2009

A general method for isolation and fractionation of chromatin into its four major components, DNA, RNA, histories, and nonhistone proteins, is described. The procedure avoids the use of strongly acidic or alkaline conditions, or the use of ionic detergents or phenol. As few as 14 × 106 cells can be used. The procedure is reasonably rapid and has been used successfully with a number of tissue culture cell lines. The chromatin components are dissociated in a 3 M NaCl – 5 M urea solution containing 2-mercaptoethanol and EDTA. The DNA and high molecular weight RNA are collected by high-speed centrifugation and DNA is separated from the RNA by means of Cs2SO4 equilibrium density centrifugation. The histones, nonhistone proteins, and low molecular weight RNA's are fractionated using DEAE-cellulose column chromatography and polyacrylamide gel electrophoresis. A small amount (< 1%) of protein is present in the DNA and RNA fractions. At least 11 low molecular weight RNA subfractions can be detected by means of polyacrylamide gel electrophoresis.
Cinnamate-4-Hydroxyiase in Polyporus hispidus
Authors: Vance C. P. , Nambudiri A. M. D. , Towers G. H. N.
Published: December 2009

A cell-free system capable of hydroxylating cinnamic acid to p-coumaric acid has been isolated from 10-day-oid cultures of Polyporus hispidus. The enzyme requires NADPH and FAD for maximum activity. Enzyme activity appears to be localized in the microsomal fraction with slight activity occurring in the mitochondrial fraction. The optimum pH for enzymatic activity is 7.5. This is the first report on any properties of this enzyme in a fungus. The enzyme differs from the known plant enzyme in its FAD requirement.
In Vitro Effects of Estrogens on the Δ4-Reduction of Testosterone by Rat Prostate and Liver Preparations
Authors: Lee D. K. H. , Young J. C. , Tamura Y. , Patterson D. C. , Bird C. E. , Clark A. F.
Published: December 2009

The inhibitory effects of six estrogens (estradiol, estriol, ethynylestradiol, mestranol, diethylstilbestrol, and chlorotrianisene) on testosterone Δ4-reduction were studied in rat prostate and liver preparations. In the prostate homogenates only those estrogens with a complete steroid structure and a free phenolic hydroxyl group at position 3 of the steroid nucleus inhibited testosterone 5α-reduction when present at 600 times the concentration of testosterone. The inhibition by estradiol was found to be competitive for prostate homogenate, microsomal, and nuclear preparations. In the liver preparations (homogenate, microsomal, and soluble fractions) all six estrogens inhibited significantly when present at the same concentration as testosterone; diethylstilbestrol and ethynylestradiol were the most effective ones.
Enzymatic Formation of Cardiolipin from Phosphatidylglycerol by the Transphosphatidylation Mechanism Catalyzed by Phospholipase D
Authors: Stanacev N. Z. , Stuhne-Sekalec L. , Domazet Z.
Published: December 2009

The enzymatic formation of cardiolipin-3H from purified phosphatidylglycerol-2′-3H catalyzed by phospholipase D is described. Some general conditions for this formation are studied and discussed. The formation of cardiolipin is independent of cytidine coenzymes and takes place by a transphosphatidylation reaction mechanism in which phosphatidylglycerol serves as the acceptor and as the donor of the phosphatidyl group.
Transient-Phase and Steady-State Kinetics for Inhibited Enzyme Systems. II. Double-Intermediate Mechanisms
Authors: Hijazi Nasrat H. , Laidler Keith J.
Published: December 2009

Equations for the pre-steady state and the steady state are derived for enzyme systems in which the enzyme E, the substrate A, and an inhibitor Q are present together, the enzyme concentration being much lower than the concentrations of A and Q. Various mechanisms are considered, ail of them involving two intermediates EA and EA′ (e.g. an acyl enzyme). When the inhibition is reversible the transient phase is followed by the establishment of a steady state. It is shown how experimental pre-steady-state and steady-state results can be analyzed to obtain rate constants, including those for the binding of inhibitor. If the binding of inhibitor is irreversible there is no steady state.
A Spin-Labelled Fluorescent Probe for Membranes
Authors: Long R. A. , Hsia J. C.
Published: December 2009

The compound N-(1-oxyl-2,2,6,6-tetramethyl-4-piperidinyl)-5-dimethylammonaphthalene-sulfonamide (DNS-SL(6)) was used as a probe to investigate the egg lecithin – cholesterol – water system. DNS-SL(6) is localized in the polar head group regions of the phospholipid bilayers and undergoes rapid anisotropic motion. Addition of cholesterol has little effect on the mobility or location of the probe but decreases the solubility of the compound in the polar head group region of the bilayers. The partition coefficient of DNS-SL(6) between egg lecithin and buffer is 5.0 × 103 at 22 °C, compared with a partition coefficient of 2.9 × 102 for egg lecithin containing 50 mol % cholesterol. The implications of the effects of cholesterol on membrane structure and passive permeability properties are discussed.
Different Chromatographic Forms of Neurospora crassa Nucleases Specific for Single-Stranded Nucleic Acids
Authors: Mills Cecily , Fraser M. J.
Published: December 2009

Three fractions with single-strand-specific nuclease activity have been isolated by chromatography on phosphocellulose and hydroxyapatite from Neurospora crassa conidia. Two of the fractions containing, respectively, 9% and 7% of the total nuclease activity recovered had the same chromatographic and enzymological properties as the endonuclease and exonuclease previously reported. The third fraction, containing 84% of the total nuclease activity recovered, was shown to contain a mixture of single-strand-specific endonuclease and exonuclease activities enzymologically indistinguishable from the activities previously described. The two components of this fraction were resolved by polyacrylamide gel electrophoresis. A fraction with the same chromatographic properties was also isolated from N. crassa mycelia in which single-strand-specific endonuclease, but no exonuclease activity, was detected. The two mycelial endonuclease fractions were enzymologically indistinguishable. The results were shown not to be due to artifacts of chromatography. Different batches of N. crassa conidia have been found to be heterogeneous with respect to contents of endonuclease and exonuclease.
Biosynthèse des glycoprotéins dans le parenchyme pulmonaire. I. Activité mannosyl-transférase dans les fractions subcellulaires des pneumocytes
Authors: Levrat Christiane , Louisot Pierre
Published: December 2009

An enzyme system exists in the membrane fraction of the alveolar cells of the lung, which catalyzes the transfer of mannose from guanosine diphosphate mannose (GDP-mannose) to an endogenous acceptor. The system is present in pellets that sediment at 10 000 g and 100 000 g. The optimal pH of the mannosyl transfer system is 5, and its optimal temperature is 27 °C. It is activated by Mn2+ and Mg2+ but is unaffected by Triton X 100, puromycin, and cycloheximide. The Michaelis constant for the substrate GDP-mannose is different for the enzyme sedimented by 10 000 g than for that by 100 000 g. Of a variety of nucleotides tested, only the guanosine nucleotides were effective as inhibitors of the activity of the two fractions.
The Effect of Vitamin A Deficiency on the Incorporation of D-Glucosaamine-1-14C into Homogenates and Subcellular Fractions of the Rat Retina
Authors: Merritt R. D. , Matuk Y.
Published: December 2009

The incorporation of D-glucosamine-1-14C into homogenates, smooth and rough microsomes, and the cell sap of retinas from vitamin-A-deficient and pair-fed vitamin-A-supplemented rats was studied. The results indicate that vitamin A deficiency produced a highly significant increase in the incorporation of D-glucosamine-1-14C into the trichloroacetic-acid-precipitable material from the different fractions studied. The possible significance of these results was discussed.