bluuebunny/crossref_metadata_2025 huggingface dataset:
A simple method for the determination of 5-vinyloxazolidine-2-thione in rapeseed meal
Authors: Szewczuk A. , Mastalerz P. , Nadwyczawski W.
Published: December 2009

5-Vinyloxazolidine-2-thione (VOT), a goitrogenic substance occurring in plants of the family Cruciferae, was found to catalyze the reduction of iodine by azide ion, the consumption of iodine being proportional to the amount of VOT. Based on these observations, a sensitive, titrimetric method of VOT determination was developed. The new method obviates the necessity of VOT extraction from the aqueous phase with organic solvents. Sensitivity and accuracy of titrimetric determination of VOT are comparable with those of spectrophotometric procedures.
m-Hydroxybenzoic acid 4-hydroxylase from Aspergillus niger
Authors: Premkumar R. , Rao P. V. Subba , Sreeleela N. S. , Vaidyanathan C. S.
Published: December 2009

m-Hydroxybenzoic acid 4-hydroxylase was isolated and partially purified from Aspergillus niger grown in presence of m-hydroxybenzoic acid. The enzyme catalyzed the stoichiometric formation of protocatechuic acid (3,4-dihydroxybenzoic acid) from m-hydroxybenzoic acid with the consumption of NADPH and molecular oxygen. The reaction proceeded best at pH 7.2 and showed a requirement for FAD.
Secretion of bile salts by intact and isolated rat livers
Authors: Subbiah M. T. , Kuksis A. , Mookerjea Sailen
Published: December 2009

Bile was collected from bile fistula rats and from the cannulated bile ducts of normal and 2-day choline-defîcient rats during 3 h of isolated liver perfusion. The bile acids were isolated as their taurine and glycine conjugates and the component acids analyzed. In comparison to the fistula bile of normal rats, the bile of the perfused livers of both choline-free and choline-supplemented animals showed increased conjugation of bile acids with glycine and a decreased cholate/chenodeoxycholate ratio. Although the volume of bile produced was similar for both groups of animals, the choline-deficient livers showed an average of 20% decrease in total bile acid production and a 20% decrease in cholate/chenodeoxycholate ratio, when compared to the acids from the bile of the perfused normal livers. By means of combined thin-layer and gas chromatographic analysis, all biles were shown to contain lithocholic, deoxycholic, chenodeoxycholic, hyodeoxycholic, ursodeoxycholic, cholic, and α- and β-muricholic acids as well as 3β,12α-dihydroxy- and 3α,12β-dihydroxy-cholanoic acids.
In vitro O2′-methylation of sugars in E. coli RNA. III. Methylation of E. coli transfer RNA by heterologous methylases in particulate-free extracts of wheat embryo
Authors: Nichols J. L. , Lane B. G.
Published: December 2009

(1) Transfer ribonucleates (tRNA) from a relaxed strain of E. coli have been methylated by homologous enzymes from E. coli and by heterologous enzymes from wheat embryo, using particulate-free extracts supplemented with [methyl-14C]-S-adenosylmethionine.(2) For both homologous and heterologous systems, the methylation of bases greatly exceeded the methylation of sugars, although the patterns of base and sugar methylation were radically different for homologous and heterologous methylations. Thus, the primary targets for homologous methylation were pyrimidine nucleosides: ribothymidine, the primary product of base methylation, and O2′-methylcytidine in the sequence CmpU, the primary product of sugar methylation. On the other hand, the primary targets for heterologous methylation were purine nucleosides: N2,N2-dimethylguanosine, the primary product of base methylation, and O2′-methylguanosine in the sequence GmpG, the primary product of sugar methylation.(3) The results of an allied study of the methylation of wheat embryo tRNA by homologous wheat embryo methylases and heterologous E. coli methylases have also been reported.
Computer-based derivation of rate equations for enzyme-catalyzed reactions. I. Effects of modifiers on kinetics of multireactant systems
Authors: Schulz Arthur R. , Fisher Donald D.
Published: December 2009

A computer-based method for the derivation of rate equations of enzyme-catalyzed reactions under steady-state assumptions is presented. This method is based on the description of the reaction mechanism in terms of a connection matrix. The utility of the method is demonstrated by applying it to complete the derivation of rate equations of multireactant enzymic mechanisms with modifiers as discussed by Henderson.
A computer program for writing the steady-state rate equation for a multisubstrate enzymic reaction
Authors: Hurst R. O.
Published: December 2009

The determinant procedure for deriving the rate equation for an enzymic mechanism has been converted into Fortran IV language for use with a digital computer. The distribution equations for three to nine enzyme species, which may interact along all possible pathways, can be determined readily, limited only by the size of the memory capacity of the computer employed. The method permits several mechanisms to be analyzed in a matter of a few minutes.
Use of 13C in biosynthetic studies. Incorporation of acetate and formate into the fungal tropolone, sepedonin
Authors: Wright J. , Smith D. G. , McInnes A. G. , Vining L. C. , Westlake D. W. S.
Published: December 2009

13C-labeled acetate and formate were administered to cultures of Sepedonium chrysospermum. The 13C-labeling patterns in sepedonin (3,6,9-trihydroxy-3-methyl-1,3,4,7-tetrahydrocyclohepta[c]pyran-7-one) were determined by measuring the relative intensities of the 13C–H satellites in the proton magnetic resonance spectra. C-1 was labeled by acetate-1-13C, and C-4, C-5, and the methyl group by acetate-2-13C. Only C-8 was labeled by formate-13C. Malonate-2-14C and succinate-2,3-14C were less efficient precursors than acetate-1-14C, acetate-2-14C, or formate-14C. The available evidence suggests that sepedonin is formed by methylation of a polyketide intermediate derived from acetate and malonate, followed by a stereospecific rearrangement and cyclization.
Ultraviolet circular dichroism studies on complexes of synthetic polynucleotides
Authors: Wolfe Fred H. , Oikawa Kimio , Kay Cyril M.
Published: December 2009

The ultraviolet circular dichroism spectra of complexes of polyadenylic acid and polyuridylic acid in the molar ratio of 1:1 and 1:2, and that of polyguanylic acid and polycytidilic acid have been examined over the wavelength range of 300–185 mμ. Increased resolution of spectra below 225 mμ has revealed several new ellipticity bands for these complexes which should be of considerable value in the proper counting of optically active electronic transitions and their assignment in these systems, and with naturally occurring RNA's.
Urease. X. A study by gel electrophoresis of its dissociation by sodium dodecyl sulfate
Authors: Blattler D. P. , Gorin George
Published: December 2009

Urease, m.w. 480 000, treated with an excess of sodium dodecyl sulfate is converted to a product of greatly increased mobility in polyacrylamide gel electrophoresis. We estimate its weight to be about 80 000. Treatment with excess thiol and detergent yielded the same product as detergent alone, indicating that the subunit or subunits do not contain polypeptide chains linked by disulfide bonds.
Phosphonic acid analogues of carbohydrate metabolites. Dihydroxypropylphosphonic acid. II. Substrate specificity of L-glycerol-3-phosphate : NAD oxidoreductase
Authors: Baer Erich , Nazir Darius J. , Basu Hemendra
Published: December 2009

The specificity of L-glycerol-3-phosphate : NAD oxidoreductase with regard to the phosphate ester bond of L-α-glycerophosphoric acid was investigated by means of L- and DL-dihydroxypropylphosphonic acid. The phosphate ester bond was found to be an essential structural requirement for enzyme activity. Neither L- nor DL-dihydroxypropylphosphonic acid has an inhibitory effect on the oxidation of L-α-glycerophosphoric acid by L-glycerol-3-phosphate : NAD oxidoreductase.
Temperature perturbation of aqueous buffered oxaloacetate solutions
Authors: Annett R. G. , Kosicki G. W.
Published: December 2009

The 215 and 260 mμ absorbance patterns of oxaloacetic acid (OAA) solutions at various pH values, subjected to temperature shifts, show a progression in both the maximum absorbance reached and the nature of the temperature effect. The levels of pyruvate, the product of OAA decarboxylation, change appreciably as well. A multiple equilibrium system is proposed to relate OAA acid dissociations, keto–enol tautomerization, and decarboxylation.
Lipid synthesis by the monoglyceride and α-glycerophosphate pathways in sheep intestine
Authors: Cunningham H. M. , Leat W. M. F.
Published: December 2009

Double-labelled monopalmitin containing 14C-glycerol and 3H-palmitic acid was used in vitro in intestinal segments and in vivo in intestinal loops of sheep to determine if triglycerides could be synthesized by both the monoglyceride and α-glycerophosphate pathways. Total glyceride synthesis in vitro by the combined pathways was highest in 87- and 120-day foetal lambs, followed in declining order by 6- to 13-day lambs, adult sheep, and adult rats (used for comparative purposes). The minimum percentage of the glycerides synthesized by the monoglyceride pathway using 1-monopalmitin as a precursor was: foetus 20, lamb 29, adult sheep 28, and rat 45. These are minimum values because appreciable hydrolysis of 1-monopalmitin occurred during incubation: 61% in rats, 68% in lambs, 71% in adult sheep, and 80% in foetal sheep. 2-Monopalmitin was more resistant to hydrolysis by intestinal segments and loops and resulted in at least 43% synthesis of glycerides by the monoglyceride pathway in segments of adult sheep intestine, compared to 26% with 1-monopalmitin.
The metabolism of d < 1.0631 lipoprotein in the rat
Authors: Buckley J. T. , Kook A. I. , Rubinstein D.
Published: December 2009

The metabolism of d < 1.063 lipoproteins of the rat was studied in perfused liver and in vivo. Perfused livers were used to label the lipoproteins in the lipid and protein moieties with palmitate-3H and leucine-14C, respectively. The lipid moiety of the isolated doubly labelled lipoprotein was removed more rapidly than the protein moiety by perfused liver. The decrease in the ratio of the lipid-3H to protein-14C in the perfusate lipoproteins occurred in three fractions of the very low density as well as the low density lipoproteins but was not accompanied by a shift of the protein among these lipoproteins. These data are consistent with an exchange of neutral lipid between the perfusate lipoproteins and the liver. The uptake and exchange of the lipid moiety of intravenously administered, doubly labelled lipoproteins apparently also occur in vivo, where the decrease in 3H/14C ratio is more rapid.Lipoprotein neutral lipid, labelled with palmitate-3H and glycerol-14C, was taken up by the perfused liver primarily in the unhydrolyzed state, as indicated by a constant 3H/14C ratio. A portion of the neutral lipid was then hydrolyzed and the resulting fatty acids were used preferentially for the synthesis of phospholipids. Unlabelled glycerol diluted the glycerol-14C in the phospholipids further increasing the 3H/14C ratio, but had no effect on this ratio in neutral lipids. It is concluded that the neutral lipid of the d < 1.063 lipoproteins can be exchanged with a small lipid pool in the liver which provides fatty acids for phospholipid synthesis.
Analysis of hormonal–dietary interactions and the role of gluconeogenesis in the regulation of tyrosine aminotransferase in rats
Authors: Mavrides C. , Lane E. A.
Published: December 2009

The extent of stimulation of rat liver tyrosine aminotransferase by Cortisol and glucagon is inversely proportional to the basal activity of the enzyme in adrenalectomized and hypophysectomized rats. The basal activity is low in animals fed a high-carbohydrate diet leading to high hormonal stimulation of the enzyme, and high in animals fed a high-protein diet leading to minimal or no hormonal stimulation. The dietary modifications of the hormonal effects have been analyzed into two independent processes. Upon short-term starvation, the enzyme activity declines rapidly in animals previously fed a high-protein diet but rises rapidly in animals previously fed a high-carbohydrate diet. Superimposing a positive (hormonal) effect on these two diametrically opposite processes results in profound modification of the hormonal effect on the enzyme activity. The experiments further suggest that the regulation of tyrosine aminotransferase is coupled with gluconeogenic activity per se rather than with a primary hormonal action at the genetic level.
Characterization of the insoluble protein fraction in the ocular lens
Authors: Lerman Sidney
Published: December 2009

The insoluble protein fraction of the ocular lens has been studied in four different species: the rat, dogfish, bovine, and human. These studies were performed in order to characterize the composition and formation of this protein in the ocular lens. On the basis of amino acid analyses, spectrophotometric determinations, thin-layer electrophoresis on Pevikon plates, and immunochemical data, it was concluded that in the rat and dogfish lens, the insoluble protein fraction derives mainly from the soluble gamma crystallin fraction. However, in the bovine and human lens, the insoluble protein fraction appears to derive mainly from the soluble alpha crystallin fraction.
A rapid method for determining the specific activity of methylene-14C citric acid in cell extracts
Authors: Hodgson B. T. , Aldous J. G. , Patterson A. H.
Published: December 2009

The microchemical spectrophotometric method for the determination of citric acid has been found to be sufficiently specific to permit its use for the measurement of specific activity, provided the labelling occurs in the noncarboxyl carbon atoms of the molecule. The simplicity of the method permits determinations to be carried out simultaneously on large numbers of samples especially if liquid-scintillation techniques are employed for the measurement of radioactivity.
Metabolism of o-hydroxy-L- and o-hydroxy-D-phenylalanine-2-14C in the rat
Authors: Petitclerc C. , Benoiton N. Leo , D'Iorio A.
Published: December 2009

o-Hydroxy-L- and o-hydroxy-D-phenylalanine-2-14C (o-tyrosine) were injected into rats, and the metabolites were separated and identified using an amino acid analyzer coupled to the flow cell of a radioactivity counter. Within 24 h, most of the label from a dose of the L-isomer had been excreted as o-hydroxyphenylacetic acid, an unidentified conjugate thereof, o-tyramine, and o-tyrosine. The D-isomer gave rise to the same metabolites, but with more of the excreted label being due to the injected material. Nembutal anesthesia altered the excretion pattern for both isomers in favor of o-tyramine at the expense of total o-hydroxyphenylacetic acid, with the effect being more pronounced for the D-isomer. Evidence was obtained for the conversion of D-o-tyrosine to L-o-tyrosine in vitro. o-Hydroxyphenylpyruvic acid was formed in vitro from D-o-tyrosine, but not from the L-isomer. Transamination of L-o-tyrosine in vitro, or incorporation into tissue proteins, could not be demonstrated.
Metabolism of organophosphorus insecticides by liver homogenates from different species
Authors: Rao S. L. N. , McKinley W. P.
Published: December 2009

The rates of disappearance of parathion, diazinon, ethion, phorate, Di-Syston (disulfoton), methyl-parathion, Guthion (aziaphosmethyl), and ronnel effected by a hepatic oxidative desulfurating system of four different species in the presence of NAD and NADP were determined. The rates of disappearance of the last three insecticides effected by the demethylating system in the presence of reduced glutathione have been determined also. The parent insecticides were determined by gas–liquid chromatography after extracting them from the incubation mixture with hexane. The conversion products formed by the oxidative system were detected by carboxylesterase inhibition on thin-layer chromatographic plates. Inhibitors were detected in extracts from the incubation mixtures of all liver homogenates with all the insecticides studied except phorate and Di-Syston. The rates of disappearance of phorate and Di-Syston were considerably higher than the rates for the other compounds with all the liver homogenates studied except female chicken liver. Methylparathion, Guthion, and ronnel were metabolized by both systems, but the demethylating system was more active for the three compounds. The rates of disappearance of the different insecticides effected by the oxidative system were considerably less when incubated with female chicken livers than with male chicken livers. These differences were not observed with the demethylating system. The rat, guinea pig, and monkey liver homogenates were all uniformly active in metabolizing the different insecticides by the oxidative system.
Fractionation of DNA on columns of poly-L-lysine supported on kieselguhr
Authors: Helleiner C. W.
Published: December 2009

Native DNA was eluted from poly-L-lysine kieselguhr columns with a shallow gradient of NaCl DNAs rich in guanine and cytosine preceded those low in these bases in the elution, and DNAs of low molecular weight preceded those of high molecular weight. Calf-thymus DNA was fractionated to give preparations differing in buoyant density.
Synthesis of 2-thio-5-carboxymethyluridine methyl ester: a component of transfer RNA
Authors: Baczynskyj L. , Biemann K. , Fleysher Maurice H. , Hall Ross H.
Published: December 2009

2-Thio-5-carboxymethyluridine methyl ester was synthesized by condensing the mercuric salt of 2-thio-5-carboxymethyluracil methyl ester with 2,3,5-tri-O-benzoyl-D-ribosyl chloride. The product is identical to a nucleoside isolated from yeast tRNA
Two possible instances of hemoglobin E Saskatoon in Manitoba
Authors: Eng Alice C. , Vella F. , Merry C. C.
Published: December 2009

During a search for hemoglobin variants in Manitoba, two unrelated patients were found who appear to possess the trait for hemoglobin E Saskatoon (β22 Glu → Lys) on the basis of peptide map analyses of the variant fractions.
The chemical constitution of lipid A from Serratia marcescens
Authors: Adams G. A. , Singh Prem Pal
Published: December 2009

The lipopolysaccharide of Serratia marcescens, under mild hydrolysis conditions (0.25 N H2SO4 for 2.5 h at 100°), yielded a lipid A containing D-glucosamine, fatty acids, acetyl and phosphate groups (approximate molar ratio 3:9:4:2), a small amount of ethanolamine, and a trace of galactosamine. Lipid A on methylation and subsequent hydrolysis yielded 3,4-di-O-methyl-D-glucosamine and 3,4,6-tri-O-methyl-D-glucosamine identified by gas–liquid chromatography as their glucitol acetates. These sugars were further identified by ninhydrin degradation to yield 2,3-di-O-methyl-L-arabinose and 2,3,5-tri-O-methyl-L-arabinose. From these findings, it was concluded that the D-glucosamine units in the lipid A were linked 1 → 6. A negative optical rotation of the lipid A suggested a β linkage. On the basis of the glycosidic linkage evidence and additional compositional data, a possible structure for the lipid A of S. marcescens is proposed and discussed in detail.
Arabinosyladenine and arabinosylhypoxanthine metabolism in murine tumor cells
Authors: LePage G. A.
Published: December 2009

The in vitro metabolism of 9-β-D-arabinofuranosyladenine (Ara-A) in TA3 ascites tumor cells was examined and compared with that of its metabolite 9-β-D-arabinofuranosylhypoxanthine (Ara-H). Some evidence for reamination of Ara-H to Ara-A was obtained. Such reamination may explain its activity as an antiviral agent. Unlike Ara-A, Ara-H was not phosphorylated and it did not inhibit DNA polymerase. Incorporation of radioactivity from Ara-A-14C or Ara-H-14C into RNA occurred, but was shown to have proceeded by a cleavage to free base and efficient reutilization of the labeled base.
Amino acid incorporation by a cell-free preparation from rat adipose tissue. Effects of fasting
Authors: Shlossberg A. H. , Hollenberg C. H.
Published: December 2009

Ribosomes were isolated from a postmitochondrial, detergent-treated supernatant fraction prepared either from adipose tissue passed through a tissue press, or from a collection of free fat cells. Optimum conditions for incorporation were established and the kinetics of the system denned with and without artificial messenger RNA. Studies were performed on ribosomes isolated from fed, 24-h fasted, and 48-h fasted animals. There was a progressive reduction in yield and activity (per milligram ribosomal RNA) of ribosomal material recovered from animals fasted up to 48 h. In the presence of poly-U, incorporation by ribosomes from fasted animals returned towards but did not reach the fed level. This return was less complete with ribosomes from 48-h than from 24-h fasted groups although as fasting proceeded, the percentage stimulation produced by poly-U progressively increased. Sucrose density gradient analysis revealed a greater proportion of light ribosome species and a smaller proportion of polysomes in fasted than in fed animals. These results suggest that fasting, in addition to producing a progressive reduction in ribosomal yield, also caused an early loss of messenger RNA or messenger function and later an alteration of ribosome function. The effect of fasting on messenger function could not be explained by alterations in ribonuclease activity.
Heat-induced disaggregation of a multifunctional enzyme complex catalyzing the first steps in pyrimidine biosynthesis in bakers' yeast
Authors: Lue Peter F. , Kaplan J. G.
Published: December 2009

A highly purified enzyme complex (from bakers' yeast), which possesses carbamylphosphate synthetase (CPSase) and aspartate transcarbamylase (ATCase) activities and a regulatory site for both activities at which feedback inhibition is exerted by UTP, was subjected to heat at 50 °C, and all three of the above parameters were followed. A treatment of 5 min was sufficient to cause disaggregation of the complex (original molecular weight 600 000) and appearance of subunits of approximately one-quarter the size of the unheated material. These subunits possessed only the ATCase activity, which was insensitive to uridine triphosphate; no component possessing CPSase activity was found in the sucrose density gradient. The ATCase was relatively resistant to heat, but there was a rapid and roughly parallel decline in the CPSase and in the sensitivity of the ATCase to inhibition by UTP. However, the residual CPSase activity, at any duration of heating, remained fully sensitive to inhibition by uridine triphosphate. CPSase activity was optimal at temperatures between 25° and 30° whereas the ATCase followed the Arrhenius law to 37 °C. Two alternative hypotheses of the structure of the enzyme complex were put forward to account for these data. Both of these hypotheses require that CPSase activity be a function of the oligomeric aggregate and not of individual subunits.
Corticosteroid metabolism and liver glycogen in fetal and newborn mice
Authors: Burton A. F. , Greenall R. M. , Turnell R. W.
Published: December 2009

Glycogen in fetal mouse liver rose from 1 to 2 mg/g on gestational day 15 to 85 mg/g just before birth, then virtually disappeared within a few hours after birth. The rise was closely associated with the appearance of steroid reductase activity in fetal liver and an abrupt drop in the rate of dehydrogenation of corticosterone to 11-dehydrocorticosterone in the fetus. Injection of a large dose of 4 mg corticosterone or cortisol into mothers on gestational day 14 increased fetal liver glycogen on day 15 up to 25 mg/g. Postnatally, a relatively high rate of corticosterone conversion continued for several weeks, falling from a peak on day 8 to one-half this value on day 14 and one-sixth on day 30, while glycogen levels rose steadily from 1 to 20 mg/g. Injection of 150 μg corticosterone increased glycogen twofold on postnatal day 14 and sixfold on day 30 over that produced on day 8 (2, 4.5, and 12 mg/g on days 8, 14, and 30, respectively). A competitive inhibitor, 11-epicorticosterone, alone increased glycogen one-third as much as an equal weight of corticosterone. It was concluded that glycogen deposition is closely related to corticosteroid metabolism involving the 11β-hydroxysteroid:NADP oxidoreductase system which is active in fetal and in early life.
Alterations in glutamate dehydrogenase of the brain of rats of various ages
Authors: Kaur Gyan , Kanungo M. S.
Published: December 2009

The activity of glutamate dehydrogenase of the brain of 6-, 22-, 52-, and 96-week-old rats was determined. The specific activity, units per milligram DNA and units per organ, increased with growth and was highest at 22 weeks. This was probably due to a higher rate of protein synthesis during growth since the total DNA content did not change during this period. The activity declined in old age. Some biogenic amines, endogenous to nervous tissue and possibly having neurotransmitter functions, such as 5-hydroxy-tryptamine (5-HT), acetylcholine (ACh), and norepinephrine (NEp), inhibited enzyme activity; epinephrine (Ep) stimulated enzyme activity at all ages. Studies on the effects of various concentrations of the transmitters showed that the concentrations of 5-HT, ACh, NEp, and Ep producing significant regulatory effects on the enzyme may be 0.048, 0.18, 0.05, and 0.025 mg/ml, respectively.
Biosynthesis of N-acetyl-L-aspartic acid in vivo and in brain homogenates
Authors: Berlinguet L. , Laliberté M.
Published: December 2009

The content of N-acetyl-L-aspartic acid in urine from pancreatectomized dogs and alloxan-diabetic rats was measured. Urinary N-acetyl-L-aspartic acid was extracted by ethyl acetate and hydrolyzed, and the aspartic acid was determined by colorimetry. The results show that in alloxan-treated rats, N-acetyl-L-aspartic acid is present in relatively high concentration in urine, while it is completely absent in the controls. In pancreatectomized dogs, the excretion of N-acetyl-L-aspartic acid in urine is much higher than in normal dogs. Furthermore, when the citric acid cycle is inhibited by the addition of fluoroacetic acid to brain homogenates, there is accumulation of radioactive N-acetyl-L-aspartic acid in the medium whereas none could be detected in controls. It is concluded that the formation of N-acetyl-L-aspartic acid in the brain can alleviate the accumulation of an excess of acetate. It is suggested that N-acetyl-L-aspartic acid may serve as a carrier of the acetyl group.
Identification of an intermediate of phloroglucinol degradation by mass spectroscopic analysis
Authors: Jamieson W. D. , Taylor A. , Mathur D. K. , Blackwood A. C.
Published: December 2009

The product of enzymatic reduction of phloroglucinol was tentatively identified as a dihydrophloroglucinol based on thin-layer chromatography and ultraviolet spectrum. Confirmatory evidence of its identity is presented by mass spectroscopic analysis.
The role of N-acetylcycloserine in the acyl transfer reactions
Authors: Whish W. J. D. , Viswanatha T.
Published: December 2009

The ability of α-N-acetylcycloserine to participate in acyl transfer reactions has been investigated. The reaction with cinnamoylimidazole yields a stable monocinnamoyl intermediate, while that with p-nitrophenyl acetate yields a derivative which undergoes facile deacylation. The kinetics of acylation of α-N-acetylcycloserine and of the deacylation of its acetyl intermediate are reported.
Rearrangement of quinonoid dihydropteridines to 7,8-dihydropteridines
Authors: Archer M. C. , Scrimgeour K. G.
Published: December 2009

The quinonoid isomer of 6,7-dimethyltetrahydropterin has been prepared in situ from the tetrahydropterin by rapid oxidation with potassium ferricyanide. Measurements of the kinetics of the rearrangement of the quinonoid dihydropterin to 7,8-dihydropterin show that the reaction is buffer-catalyzed. Both the acidic and basic forms of the buffer participate as catalysts, with the relative contribution of each species toward the rate of the reaction being dependent on the pKa value of each buffer. Isotopic experiments have shown that the cleavage of the C—H bond at position 6 is rate-limiting during both the acid- and the base-catalyzed rearrangements.
Studies of the binding of adenine to adenine phosphoribosyltransferase
Authors: Gadd R. E. A. , Henderson J. Frank
Published: December 2009

The groups on adenine phosphoribosyltransferase and on adenine which are involved in the binding of this substrate were investigated by comparing the inhibition constant of adenine with those obtained for analogues of adenine, by determining the effect of pH on the Michaelis constant of adenine, and by protection by adenine against enzyme inactivation by N-bromosuccinimide. Those moieties of adenine which are probably involved in complexing with the enzyme include the 6-amino group, and the 3-nitrogen and the 7-nitrogen atoms. The first probably complexes as an electron acceptor and the last two as electron donors. There is probably a lack of bulk tolerance at the 6-amino, 2-carbon, and the 3-, 7-, and 9-nitrogen positions. The group on the enzyme which ionizes at pH 10.25 was tentatively identified as a phenolic hydroxyl group of tyrosine which complexes with either the 3- or 7-nitrogen atoms of adenine.
The role of magnesium ion in the adenine phosphoribosyltransferase reaction
Authors: Gadd R. E. A. , Henderson J. Frank
Published: December 2009

Low concentrations of magnesium ion stimulate, and high concentrations inhibit, the adenine phosphoribosyltransferase reaction. The stability constant of the Mg–pyrophosphoryl moiety of phosphoribosylpyrophosphate was assumed to be the same as that of Mg–adenosine triphosphate; the stability constant of the Mg–phosphoryl moiety of phosphoribosylpyrophosphate was assumed to be the same as that of Mg–adenosine monophosphate, and the concentration of the monomagnesium complex of phosphoribosylpyrophosphate was calculated as a function of magnesium ion and phosphoribosylpyrophosphate concentrations. The good correspondence between these curves and those of initial velocity under the same conditions suggest that the monomagnesium complex of phosphoribosylpyrophosphate is the true substrate of this reaction.
Multiple molecular forms of avian aldolases. IV. Purification and properties of chicken (Gallus domestkus) brain aldolase
Authors: Marquardt Ronald R.
Published: December 2009

Aldolase (fructose-1,6-diphosphate D-glyceraldehyde-3-phosphate-lyase, EC 4.1.2.13) was purified from chicken (Gallus domesticus) brain tissue. The enzyme was shown to be homogeneous according to the following criteria: purification to a constant specific activity following sequential chromatography on DEAE and Sephadex, sedimentation velocity analysis, and electrophoresis on cellulose acetate strips.Several properties of the enzyme were determined including the Stokes radius (47 Å), diffusion constant (D020 w = 4.6 × 10−7 cm2/s), sedimentation coefficient (s020 w = 8.0), and molecular weight (155 000). The enzyme has a broad pH optimum centered around 7.2. The apparent Michaelis constants for fructose 1,6-diphosphate and fructose 1-phosphate were 7 × 10−5 M and 3 × 10−2 M, respectively. The activity ratio with the above two substrates was 30.Many of the molecular properties of this enzyme are similar to those of the rabbit brain enzyme and the muscle enzymes from both chickens and rabbits. The enzymic properties of chicken brain aldolase correspond more closely to those of the rabbit brain enzyme than they do to chicken breast muscle aldolase. The amino acid composition of chicken brain aldolase was found to be quite different from chicken breast muscle aldolase with respect to certain amino acids (methionine, cysteine, tryptophan, histidine, proline, aspartate, valine, and phenylalanine).
Enzymes involved in the metabolism of thiosulfate by Thiobacillus thioparus. I. Survey of enzymes and properties of sulfite : cytochrome c oxidoreductase
Authors: Lyric Ronald M. , Suzuki Isamu
Published: December 2009

Enzymes concerned with the oxidation of thiosulfate were investigated in extracts of Thiobacillus thioparus. The organism possessed sulfite oxidase as well as adenosine-5′-phosphosulfate reductase and thiosulfate-oxidizing enzyme. Sulfite oxidase was purified 160-fold and the properties were studied. The enzyme had a molecular weight of 54 000 and one non-heme iron. The pH had a marked effect on reaction velocity and Km for sulfite, and the pK values for free enzyme and enzyme–sulfite complex were determined as 8.9 and 6.2, respectively. Chloride inhibition was noncompetitive and phosphate was uncompetitive with respect to sulfite. In many properties the T. thioparus enzyme was similar to the enzyme isolated from Thiobacillus novellus.
Enzymes involved in the metabolism of thiosulfate by Thiobacillus thioparus. II. Properties of adenosine-5′-phosphosulfate reductase
Authors: Lyric Ronald M. , Suzuki Isamu
Published: December 2009

Adenosine-5′-phosphosulfate (APS) reductase was purified from Thiobacillus thioparus extracts 25- to 46-fold and the properties were studied. The molecular weight was 170 000 and the enzyme had 1 mole of FAD, 8–10 moles of iron, and 4–5 moles of labile sulfide. Cytochrome c as well as ferricyanide served as the electron acceptor. The pH optimum shifted from 7.4 to 9.5 when cytochrome c was used instead of ferricyanide. The Km values for sulfite and AMP were reduced from 2.5 mM and 100 μM to 17 μM and 2.5 μM, respectively, with cytochrome c as electron acceptor. Properties of the T. thioparus enzyme were compared to those of APS reductase isolated from Thiobacillus denitrificans and Desulfovibrio desulfuricans.
The inhibition of chymotrypsin A4 with a homologous series of chloromethyl ketone reagents
Authors: Stevenson Kenneth J. , Smillie Lawrence B.
Published: December 2009

The inactivation of chymotrypsin A4 (CHT-A4) by a homologous phenylalkyl series of bifunctional reagents, viz. phenyl chloromethyl ketone (PCK), benzyl chloromethyl ketone (BCK), and β-phenylethyl chloromethyl ketone (βPECK) (C6H5 [CH2]n COCH2Cl where n = 0, 1, or 2), has been investigated. The inactivation of CHT-A4 by PCK has been shown to be essentially pH independent whereas inactivation by BCK and βPECK appears to be dependent on a basic group of pK 6.0–6.3. PCK has been shown to inhibit CHT-A4 by virtue of the complete S-alkylation of methionine-192. BCK and βPECK, on the other hand, inactivate CHT-A4 through a combination of partial S-alkylation of methionine-192 (0.2 and 0.3 residue, respectively) and partial alkylation of histidine-57 (0.2 and 0.4 residue, respectively). Identification of the particular residue alkylated was obtained through the isolation and analysis of alkylated and oxidized peptic peptides obtained from the alkylated CHT-A4 by the diagonal peptide ionophoresis technique.The initial site of alkylation of BCK and βPECK on the imidazole ring of histidine-57 has not been identified. However, a unique histidine derivative has been isolated from the acid hydrolysate of oxidized histidine-57 peptide from CHT-A4–βPECK and is suggested to be 2(or 4)-hydroxymethylhistidine.On the basis of the present studies on the phenylalkyl series and on studies reported earlier on a phenylalkylamido halomethyl ketone series of bifunctional reagents which alkylates only methionine in CHT-A4, it has been suggested that alkylation of histidine by members of the phenylalkyl series is primarily a result of binding to the hydrophobic binding site of CHT-A4 rather than binding to the acylamido binding site as is suggested to be the case for the phenylalkylamido series.
CMorophyllides c
Authors: Wasley J. W. F. , Scott W. T. , Holt A. S.
Published: December 2009

"Chlorophyllides" c were obtained from methanol–tetrahydrofuran extracts of Phaeodactylum tricornutum by retention on an anion exchange resin. They were esterified (diazomethane) and separated by adsorption chromatography on powdered sucrose. The less adsorbed fraction was designated c1; the other c2. Both pigments possess two main visible absorption bands at ca. 629 and 582 nm. The ratios of the intensities of absorption in dioxane of band I to band II were 1.09 and 0.69, respectively.Treatment with methanolic hydrogen chloride yielded the respective chloroporphyrin e6 trimethyl esters, whose molecular weights (determined by mass spectrometry) were 634 and 632. The n.m.r. spectra showed the presence of the C7 acrylate group. Since both c1 and c2 yielded pyrroporphyrin XV, the structure of c2 requires C2 and C4 vinyl groups and that of c1 a vinyl group at either C2 or C4.Extinction coefficients of the phyllins and of the magnesium-free derivatives in various solvents were determined. Equations for the measurement of chlorophyllide c concentration in the presence of chlorophylls a and b are given.
Studies on the control of fatty acid oxidation in liver preparations from chick embryos
Authors: Koerker D. J. , Fritz I. B.
Published: December 2009

The characteristics and developmental pattern of the metabolic pathway for fatty acid oxidation were investigated in liver slices and mitochondria prepared from chick embryos of varying ages. In 8-day-old chick embryos, hepatic fatty acid oxidation was readily measurable. The incorporation of labelled palmitate into CO2 was increased twofold by carnitine in liver slices of 8-day-old chick embryos but by nearly sixfold to tenfold in tissues prepared from 10- or 12-day-old embryos. A similar increase was seen in the degree of augmentation of ketogenesis induced by carnitine in liver slices prepared from the 10-day-old embryo, suggesting an increased carnitine palmitoyltransferase activity in liver cells during the stage of development from 8 to 10 days. Palmitoyl-CoA was not metabolized in the absence of carnitine, whereas the palmitoyl portion of palmitoylcarnitine readily supported respiration by embryonic chick liver mitochondria. In the presence of adequate amounts of albumin, good respiratory control was evident.The administration of glucose to chick eggs which had previously been incubated for approximately 4.5 days resulted in changes in the metabolism of embryos killed 5 days later, which indicated that tissues of the chick embryo were capable of integrative metabolic adaptations in response to changes in substrate supply.
Large-scale preparation and some properties of penicillopepsin, the acid proteinase of Penicillium janthinellum
Authors: Sodek J. , Hofmann T.
Published: December 2009

The acid proteinase penicillopepsin has been isolated from the culture filtrate of the mould Penicillium janthinellum, grown in submerged culture in a 1500 l fermentor. After concentrating the enzyme by adsorption on diethylaminoethylcellulose, it was purified by chromatography on aminoethylcellulose and phosphocellulose. The yield of the purified enzyme was about 2.6 g (37% recovery). It was shown to be homogeneous on acrylamide gel. Crystals suitable for X-ray analysis were prepared from the pure enzyme. Both the N- and the C-terminal amino acids appear to be alanine, although N-terminal serine has also been found in varying amounts. The purified enzyme is stable to prolonged exposure in the pH range 2.2–6.0, and at pH 4.9 retains 100% activity for 1 h at 55°. The crystals retain full activity at room temperature for many weeks.
Carbon dioxide fixing pathways of glutamic acid synthesis in the rumen
Authors: Milligan L. P.
Published: December 2009

Rumen contents were incubated with NaHCO3-14C and the pattern of incorporation of 14C into glutamic acid was determined. Label was found in C-1, C-2, and C-5 of glutamic acid. This distribution pattern suggested the simultaneous occurrence of the forward tricarboxylic acid (TCA), the atypical forward TCA, and the reverse TCA pathways of glutamate synthesis in the mixed rumen population; these pathways were estimated to account for 63%, 9%, and 28%, respectively, of the glutamate synthesized by pathways entailing CO2 fixation. The contributions of the individual pathways were not influenced by the ration of the host. The significance of these alternate pathways of glutamate synthesis in the metabolism of rumen microorganisms is discussed.
Effects of chloramphenicol and its isomers and analogues on the mitochondrial respiratory chain
Authors: Freeman K. B.
Published: December 2009

The nature of the inhibition of mitochondrial respiration by chloramphenicol isomers and analogues has been examined further to explain differences in their action in several systems. With coupled rat-liver mitochondria, D-threo-chloramphenicol inhibited β-hydroxybutyrate oxidase 50% at 1.3 mM and succinoxidase at 3.5 mM. L-threo-Chloramphenicol inhibited 50% at 3.6 mM and 3.7 mM, respectively. The sensitivity of succinoxidase to D-chloramphenicol was lost on freezing the mitochondria. Chloramphenicol inhibited dinitrophenol-activated ATPase with 50% inhibition at 6 mM for the D-isomer and at 2.2 mM for the L isomer. The inhibition of NADH oxidase of frozen–thawed beef-heart mitochondria by D-chloramphenicol analogues was dependent on the analogues' solubility properties. A relationship which approximates a straight line was obtained by plotting log (1/concentration50% inhibition) against log (solubilityethanol/solubilitywater) indicating that the inhibition was greater for analogues which would partition into an organic phase. The inhibition did not depend on the orientation of the mitochondrial membrane; the NADH oxidase of submitochondrial particles prepared either by sonication or with digitonin had the same sensitivity to the analogues. The inhibition of NADH oxidase by rotenone and amytal fitted into the straight line relationship obtained with the chloramphenicol analogues. Beef-liver glutamic dehydrogenase was inhibited by some of the analogues of chloramphenicol with the same relationship as for the inhibition of NADH oxidase. It was suggested that the inhibition of NADH oxidase and glutamic dehydrogenase depends on the inhibitors reacting with a hydrophobic site on the enzyme.
Studies on the aminoxyalanine methyl ester and its α-N-acetyl derivative
Authors: Whish W. J. D. , Viswanatha T.
Published: December 2009

The ability of the aminoxyalanine methyl ester and its α-N-acetyl derivative to promote the hydrolysis of cinnamoylimidazole was found to increase with time. The time-dependent increase in the activity could not be explained on the basis of the hydrolysis of the functional groups or the cyclization to yield the parent isoxazolidone ring. The formation of an active derivative in the solutions of these compounds is suggested.
Reduction potentials of tetrahydropterins
Authors: Archer M. C. , Scrimgeour K. G.
Published: December 2009

The standard reduction potential for the quinonoid-dihydropterin–tetrahydropterin couple has been determined by polarography and by reduction of a series of redox indicators. E0 for the couple is + 0.60 V and E0′ is+ 0.15 V.
Deoxyribonucleic acid polymerase from nuclei of rat intestinal mucosa
Authors: Leung F. Y. T. , Zbarsky S. H.
Published: December 2009

An extract with DNA polymerase activity was prepared from nuclei of intestinal mucosa of the rat. Chromatography of the crude extract on DEAE-cellulose yielded three enzymically active fractions: I, II, and III. Each fraction could be resolved further into two components with DNA polymerase activity by rechromatography on smaller columns of DEAE-cellulose. A similar result was obtained by gel filtration of fraction II material on Sephadex G-150. The result of sucrose density gradient centrifugation of the fractions obtained by gel filtration suggested that each still consisted of a mixture of proteins with DNA polymerase activity. The approximate molecular weights of the active proteins, estimated by comparison with marker proteins, ranged from 25 000 to 300 000. Partially purified DNA polymerase (fraction II) required for activity the four deoxyribonucleoside triphosphates, Mg2+, 2-mercaptoethanol, and DNA template. The optimum pH for activity was 8.0 in Tris–HCl buffer and 7.4 in phosphate buffer. The two components obtained by gel filtration of fraction II differed in their requirements for DNA template. The one of smaller molecular size was more active with native DNA whereas the larger was equally active with either native or heat-denatured DNA.
Palmitic acid-1-14C oxidation by skeletal muscle mitochondria of dystrophic mice
Authors: Lin C. H. , Hudson A. J. , Strickland K. P.
Published: December 2009

Cofactor requirements for the oxidation of palmitate-1-14C by 600 × g supernatant fraction of mouse skeletal muscle homogenate and by skeletal muscle mitochondria are described. Optimal oxidation of palmitate-1-14C by skeletal muscle mitochondria requires the presence of carnitine, ATP, CoA, and a Krebs cycle intermediate (e.g. succinate). Succinate, malate, alpha-ketoglutarate, and oxaloacetate are all equally effective in supporting the oxidation, but isocitrate is less effective. The oxidation of palmitate-1-14C by 600 × g supernatant fraction of muscle homogenate as well as by skeletal muscle mitochondria from dystrophic mice is significantly decreased compared with that of the normal littermate controls. The present results, together with the previous findings, suggest that the decrease in oxidation of palmitate-1-14C by the dystrophic muscle preparations is most likely due to a defect in one or more of the steps of the Krebs cycle.
Kinetic studies of sulfite; cytochrome c oxidoreductase, thiosulfate-oxidizing enzyme, and adenostne-5′-phosphosulfate reductase from Thiobacillus thioparus
Authors: Lyric Ronald M. , Suzuki Isamu
Published: December 2009

Kinetic studies were carried out on three enzymes purified from Thiobacillus thioparus: sulfite: cytochrome c oxidoreductase, thiosulfate-oxidizing enzyme, and adenosine-5′-phosphosulfate reductase. From the initial velocity and product inhibition studies a tentative kinetic mechanism was proposed for each enzyme reaction.
Partial purification and properties of L-phenylalanine ammonia-lyase from Streptomyces verticillatus
Authors: Emes A. V. , Vining L. C.
Published: December 2009

Phenylalanine ammonia-lyase (EC 4.3.1.5) was purified 40-fold from a cell homogenate of Streptomyces verticillatus. In many respects the enzyme was similar to phenylalanine ammonia-lyases isolated from plants and fungi. It was most active at pH 9.0 and the Km for L-phenylalanine was 1.6 × 10−4 M. It showed no requirement for metal ions but was inhibited by heavy metals, some sulfhydryl reagents, and carbonyl reagents. The Stokes' radius was estimated by gel filtration to be 5.45 nm. Sucrose gradient centrifugation gave an s20,w of 10.0, leading to calculated values of 226 000 and 1.61 for the molecular weight and frictional ratio, respectively, if a partial specific volume of 0.725 ml/g is assumed. The enzyme deaminated o-, m-, and p-fluoro-, p-chloro-, and p-methyl-phenylalanine but was without action on L-tyrosine. It was inhibited by trans-cinnamic acid and certain phenylalanine derivatives, as well as by some less closely related aromatic compounds, but not by trans-cinnamamide.
Accumulation of dehydrofusaric acid and its conversion to fusaric and 10-hydroxyfusaric acids in cultures of Gibberella fujikuroi
Authors: Pitel D. W. , Vining L. C.
Published: December 2009

Gibberella fujikuroi (Saw.) Wr. strain 917 produced dehydrofusaric acid in high yield when cultured in a defined medium containing adequate nitrogen. The metabolite was separated by partition chromatography from the small amount of fusaric acid present. Fusaric and dehydrofusaric acids can be interconverted by the culture since radioactivity administered as either 14C-labeled fusaric acid or 14C-labeled dehydrofusaric acid was distributed into both metabolites. Radioactivity also accumulated in more polar substances. These were isolated from cultures administered acetate-2-14C and the main radioactive product was identified as 10-hydroxyfusaric acid. 11-Chloro-10-hydroxyfusaric acid was tentatively identified in the culture extract, but may be an artifact.
Urease: the 12 S form
Authors: Lynn K. R.
Published: December 2009

Ureolytically active urease with a sedimentation coefficient of 12 S has been observed as a major component following centrifugation in sucrose gradients prepared with citrate buffers of high ionic strengths.