bluuebunny/crossref_metadata_2025 huggingface dataset:
On the mechanism of messenger RNA binding by E. coli 70 S ribosomes
Authors: Krahn P. M. , Paranchych W.
Published: December 2009

Various aspects governing the interaction of 3H-poly U with E. coli ribosomes have been investigated with the use of a simple filtration technique. The binding of poly U to ribosomes was found to be temperature-dependent; i.e. the polynucleotide attaches most efficiently at a critical temperature consistent with its ordered-state to random-coil transition. Moreover, the interaction of poly U with ribosomes was shown to be magnesium-dependent at all temperatures examined, although the interaction was markedly stimulated by high magnesium concentrations (greater than 12 mM) at 0°. At 23° no increase in association of poly U with ribosomes was noted above 12 mM magnesium. Ribosome – poly U complexes exhibited reduced association, analogous to polynucleotide "melting" profiles, and yielded a Tm of 25° (the temperature at which the complex is half dissociated). The decreased association of poly U with ribosomes at elevated temperatures, in addition to other observed effects in the presence of the two protein perturbant solvents ethylene glycol and dimethyl sulfoxide, has led to the conclusion that hydrogen bonding probably plays a prominent role in the stabilization of the ribosome–mRNA complex.
The effect of cell-sap fractions on incorporation of amino acids into protein in rat liver cell-free systems
Authors: Liew C. C. , Korner A.
Published: December 2009

The ability of rat liver cell sap, pH 5 enzyme preparation, partially purified transfer factors I and II (PTF), and a mixture of the latter two to support protein synthesis in a ribosome system in vitro was compared. Protein synthesis increased as the ratio of pH 5 enzyme to ribosomes and PTF was raised and as the ratio of PTF to pH 5 enzymes and ribosomes was raised. The best ratios of these fractions were not as efficient as unfractionated cell sap at supporting protein synthesis; some factor had been lost or inactivated during fractionation. Incubation with cell sap caused changes in ribosomal profiles leading to less polysomes and more monomeric and dimeric ribosomes but no changes were noted on incubation with pH 5 enzymes and PTF.
The sulfhydryl groups of muscle phosphorylase. V. The reactive sulfhydryl peptides
Authors: Zarkadas C. G. , Smillie L. B. , Madsen N. B.
Published: December 2009

Skeletal muscle glycogen phosphorylase a and b have previously been shown to consist of four and two subunits, respectively, each having a molecular weight of 92 500. Our studies have indicated that there is a minimum of eight and a maximum of nine unique cysteine sequences in the enzyme providing additional evidence that the subunits are identical. From the known sequences of the nine sulfhydryl peptides it was possible to isolate and characterize the reactive sulfhydryl peptides of phosphorylase. Careful column chromatographic purification of alkylated peptides derived from peptic digestion showed that there are two cysteine residues per monomer of phosphorylase b whose rate of reaction with iodoacetamide approaches that of model compounds and whose alkylation does not affect significantly the enzymic properties of the protein. These two cysteines have been identified in sequences corresponding to numbers 2 and 5 previously elucidated. These sequences have been extended in the present work and may now be written as: No. 2 Asn–Gln–Lys–Ile–CMC–Gly–Gly–Try–Gln–Ser, and No. 5 Gly–CMC–Arg–Asp–Pro–Val–Arg–Thr–Asn–Phe.
Reduction of nonheme iron in the respiratory chain of Escherichia coli
Authors: Bragg P. D.
Published: December 2009

A particulate fraction of E. coli contained flavoprotein, nonheme iron, ubiquinone, and cytochromes b1 and o. About 20% of the nonheme iron was reducible by NADH, ascorbate – phenazine methosulfate, and dithionite to a form which reacted rapidly with o-phenanthroline. Reduction of this o-phenanthroline-reacting nonheme iron species by NADH was inhibited by 2-heptyl-4-hydroxyquinoline N-oxide which inhibited the respiratory chain between ubiquinone and cytochrome b1. Reduction of cytochrome b1 could occur without reduction of this nonheme iron species. These results are discussed relative to the position of nonheme iron in the respiratory chain of E. coli.
The conformation of an atypical IgG myeloma protein and its papain fragments
Authors: Connell G. E. , Dorrington K. J. , Lewis A. F. , Parr D. M.
Published: December 2009

An immunoglobulin IgG (Sackfield) which is known to have polypeptide chains shorter than those of typical proteins of its class has been subjected to fragmentation by papain in the presence of cysteine. One fragment was recovered which was indistinguishable from normal Fc fragment. The other fragment was related to normal Fab fragment but differed from it in several of its properties. The molecular weight was only one-half that of normal Fab. The optical rotatory dispersion spectrum of IgG (Sackfield) had features which differed from those of typical IgG myeloma proteins. The optical rotatory dispersion spectrum of Fc (Sackfield) was identical with those of other myeloma proteins, while the Fab (Sackfield) spectrum reflected the differences observed in the parent protein.
The effect of chloramphenicol on the induction of cyanase In Escherichia cott
Authors: Taussig A. , Ronnen Eva
Published: December 2009

The early stages of cyanase induction were studied. It was found that 30 s elapsed at 37 °C and 60 s elapsed at 22 °C before cyanase-forming capacity of Escherichia coli could be demonstrated. This period was termed inducer-dependent stage (i.d.s.). An additional period of 120 s at 22 °C was needed for demonstration of enzyme synthesis. This period was termed inducer-independent stage (i.i.s.). It was found that in contrast to β-galactosidase, the i.d.s. as well as the i.i.s. were inhibited by chloramphenicol. Pulse-labelling experiments showed an increase in m-RNA obtained from induced bacteria in the presence of chloramphenicol compared with noninduced bacteria treated with chloramphenicol, indicating that chloramphenicol does not interfere with transcription. However, the m-RNA formed in the presence of chloramphenicol did not participate in translation upon removal of the antibiotic. This evidence, in addition to the time course of the inhibition, suggests that the inhibition of induction is caused by chloramphenicol preventing the attachment of the ribosomes to cyanase-specific m-RNA.
Effect of hypophysectomy on amino acid, thymidine, and orotic acid incorporation into the mucosa of the small bowel
Authors: Levitan Reuven , Havivi Eli
Published: December 2009

After hypophysectomy, the incorporation of tritiated thymidine and orotic acid into DNA and RNA, respectively, decreased in the intestinal mucosa. Hypophysectomy lowered the incorporation of amino acids into protein in vitro, yet the proportion of polysomes remained unaltered. After hypophysectomy, the polysomes were impaired in their ability to incorporate amino acids in vitro.
A preliminary survey of the effects of some antibiotics on the incorporation of amino acids by a nuclear fraction of young chick blastodiscs
Authors: Trevithick J. R. , Wainwright S. D.
Published: December 2009

A "nuclear fraction" from young chick blastodiscs incorporated lysine and leucine into polypeptide in vitro. The system was only slightly inhibited by high concentrations of cycloheximide, puromycin, and nuclease enzymes, but was markedly inhibited by chloramphenicol and dinitrophenol. Protein extracted by 0.2 M HCl was labelled by lysine in vitro.
Studies on the biochemical properties of surface component of normal and SV-40 transformed 3T3 mouse cells
Authors: Sheinin Rose , Onodera Kazukiyo
Published: December 2009

A specifically marked surface component from 3T3 mouse cells and from SV-40 transformed 3T3 cells has been isolated and partially purified. These components have been shown to be free of RNA, DNA, and lipid. They do contain carbohydrate and peptide, probably as glycoprotein.
In vitro synthesis of β-galactosidase by membrane fractions of Escherichia coli
Authors: Chefurka W. , Yapo A. , Nisman B.
Published: December 2009

The induction of β-galactosidase by a membranous fraction P1, prepared by digitonin lysis of spheroplasts of Escherichia coli, was studied in vitro. Electron micrographs of P1 show it to be a heterogeneous mixture of smooth vesicles, rough vesicles, rough vesicles attached to DNA, and ribosomes attached to DNA. P1 was subfractionated by differential centrifugation into an active heavy fraction, P4, and a relatively inactive light fraction, Pm. The P4 fraction consisted mainly of rough vesicles while the Pm fraction consisted mainly of smooth vesicles, but also of some rough vesicles. These vesicles of the Pm fraction were further separated by discontinuous sucrose gradient centrifugation.The induction of β-galactosidase by P1, P4, and Pm fractions was not related to mild contamination by unbroken viable spheroplasts. It was only partially sensitive to DNase and RNase. High concentrations of actinomycin D were required for complete inhibition of activity. This suggests that the transcription and translation components were shielded by the membranes. The synthetic activity of Pm was enhanced by fortification with DNA and/or S30. Only lac-containing DNA was active. The induction of β-galactosidase by this reconstituted system showed an absolute requirement for Pm membranes and for the inducer but only a partial requirement for nucleoside triphosphates. It was completely inhibited by puromycin and chloramphenicol.
Hemoglobin Broussais: α90 Lys → Asn
Authors: Vella F. , Charlesworth Deborah , Lorkin P. A. , Lehmann H.
Published: December 2009

A hemoglobin with the electrophoretic properties of hemoglobin K or J was found in three generations of a Canadian family in Saskatoon. By analysis of the tryptic peptides of the whole globin and of the isolated abnormal α chains, it was characterized as α90 Lys → Asn (hemoglobin Broussais). This is the second report of the occurrence of this hemoglobin variant.
Reactions of superoxide anion, catechols, and cytochrome c
Authors: Miller Richard W.
Published: December 2009

Photoreduced flavins react very rapidly with molecular oxygen to give the superoxide anion. This radical is highly reactive with cytochrome c and p-nitro blue tetrazolium salts. The reduction of these electron acceptors under aerobic conditions is strongly inhibited by catechols such as 1,2-dihydroxybenzene-3,5-disulfonate (Tiron) and 3,4-dihydroxybenzoic acid. Tiron also is essential for the photosensitized reoxidation of ferrocytochrome c in the presence of flavins and oxygen. These findings support a proposal that catechols react rapidly with superoxide anion to yield a new oxidizing species. This fast reaction effectively prevents the oxygen radical from participating in slower oxidation–reduction processes. Inhibition by Tiron of the oxygen-dependent catalytic reduction of electron acceptors is therefore indicative of the participation of enzymatically generated superoxide anion. The proposed explanation of the observed phenomena does not require interaction of the catechol inhibitor with the iron–sulfur chromophores of superoxide-forming metalloflavoproteins.
Agents affecting lipid metabolism. XXXIX. Effect of combined administration of ethyl chlorophenoxyisobutyrate and cholestyramine on cholesterol biosynthesis in the rat
Authors: Cayen M. N. , Dvornik D.
Published: December 2009

Rats were fed ethyl p-chlorophenoxyisobutyrate (CPIB) and cholestyramine, alone and in combination. Regarding levels of circulating cholesterol, cholestyramine had no effect, while a fall was observed with CPIB given alone or in combination with cholestyramine. Subsequently, the combined effect of both agents was elicited by measuring the incorporation of 2-14C-acetate into cholesterol by liver homogenates of treated rats; addition of CPIB decreased the cholestyramine-induced increase in the rate of hepatic cholesterol synthesis.
The effect of urea upon the activity measurement of cod muscle cathepsin with hemoglobin substrate
Authors: Wojtowicz M. B. , Odense Paul
Published: December 2009

The effect of urea upon the activity of cod muscle cathepsin on hemoglobin substrate was studied.The apparent activation of the enzyme, found when urea was present, either added to the hemoglobin substrate before incubation or added at the start of the enzyme incubation with substrate, may be explained as an increase in the solubility of the enzyme reaction products in the presence of urea when trichloroacetic acid is added to end the incubation. This was shown by addition of urea together with trichloroacetic acid at the end of incubation. This resulted in activity measurements higher than any when urea was present during the incubation, and demonstrated that the effect of urea is to inactivate rather than activate the enzyme.
Hemoglobin Mahidol: a new hemoglobin α-chain mutant
Authors: Pootrakul S. , Dixon G. H.
Published: December 2009

A slow (less anionic) hemoglobin mutant has been detected by starch gel electrophoresis of hemoglobin from three unrelated patients in Bangkok. Dissociation of the abnormal hemoglobin with p-hydroxymercuribenzoate showed that the α-chain was the site of the mutation. The mutant α-chain was isolated by carboxymethylcellulose chromatography in 8 M urea and 0.05 M β-mercaptoethanol. Peptide maps of trypsin and cyanogen bromide cleaved α-chain indicated that the amino acid alteration of the mutant was in the peptide corresponding to residues 62–76 of the α-chain. Further cleavage of this peptide with 0.25 M acetic acid at 110 °C showed that residue 74 was changed from an aspartyl to a histidyl residue, a mutation not previously described. It is proposed that this new hemoglobin α274His β2A be called hemoglobin Mahidol after Mahidol University in Bangkok. In one of the three patients showing hemoglobin Mahidol, interaction with α-thalassemia occurs and, in this patient, hemoglobin A is totally absent, being replaced by hemoglobin Mahidol together with some hemoglobin H (β4A).
Studies on the mitochondrial adenosine triphosphatase system. VI. Coupling activity of F1 subunits
Authors: MacLennan David H. , Paulson Carlton W.
Published: December 2009

In the presence of subsaturating levels of coupling factor I (F1 and saturating levels of the oligomycin-sensitivity-conferring protein (OSCP), the addition of subunits of F1 enhanced the rate of ATP-32Pi exchange, the rate of ATP-energized DPN+ reduction by succinate, and P/O ratios in submitochondrial particles prepared by ammonium extraction (A particles). It is postulated that a limited number of ultrastructural lesions in A particles results in uncoupling. The reconstitution of A particles by the different coupling proteins could result from the repair of several types of lesions in the membrane.
The effect of puromycin and aminonucleoside on kidney proteins
Authors: Nicholls D. M. , Westall C. G. , Aung B. D.
Published: December 2009

The incorporation of 14C-leucine into microsomal and soluble protein was measured in postmitochondrial supernatant of kidney homogenates prepared from rats receiving puromycin or aminonucleoside. No short-term effects were observed in kidney. When puromycin or aminonucleoside, 7.5 μmoles or 5.0 μmoles, respectively, was administered daily for 11 days, an increased incorporation was observed with both substances but the increase was considerably greater with aminonucleoside. In kidney preparations from aminonucleoside nephrotic rats, the incorporation of 14C-leucine was increased in both albumins and globulins, but to a larger extent in the albumins of the soluble protein fraction.
The 3′-hydroxyl termini in yeast ribosomal RNA
Authors: Olver K. M. , Lane B. G.
Published: December 2009

(1) A fraction of high molecular weight RNA was prepared by subjecting total yeast RNA to selective precipitation from aqueous 2.5 M sodium chloride solution at 0°. This "insoluble" RNA accounted for about 80% of the total yeast RNA.(2) The "insoluble" RNA and various of its subfractions obtained by preparative sucrose density-gradient centrifugation were subjected to electrophoretic and end-group analyses.(3) One subfraction, largely composed of 17 S RNA from the smaller ribosomal subunit, was found to contain adenosine as the principal 3′-hydroxyl terminal nucleoside residue. Based on the analysis of 3′-hydroxyl termini, RNA in this subfraction had a mean residue length of about 1600 nucleotides.(4) Another subfraction, largely composed of 26 S RNA from the larger ribosomal subunit, was found to contain uridine as the principal 3′-hydroxyl terminal nucleoside residue. Based on the analysis of 3′-hydroxyl termini, RNA in this subfraction had a mean residue length of about 2400 nucleotides.(5) Electrophoretic and end-group analyses of other subfractions have been reported and discussed in terms of the possible origin of the RNA in these fractions, and also in terms of the way in which the RNA of these fractions might tend to affect the end-group analysis of both the parent fraction and the principal subfractions, which contain RNA from the ribosomal subunits.
Effect of N-methylation and chain length on kinetic constants of trypsin substrates. ε-N-Methyllysine and homolysine derivatives as substrates
Authors: Seely John H. , Benoiton N. Leo
Published: December 2009

The action of trypsin on the following amino acid derivatives has been investigated: the ethyl esters of ε-N-mono-, ε-N-di-, and ε-N-tri-methyl-L-lysine; the ethyl esters of the homologues of lysine and arginine; the methyl ester and amide of the α-N-benzoyl-DL-homolysine; the methyl esters and amides of the α-N-benzoyl derivatives of ε-N-di- and ε-N-tri-methyllysine; and poly ε-N-methyllysine. Derivatives of L-ornithine, DL-2,8-diaminooctanoic acid, ε-N-dimethyl-, ε-N-trimethyl-, and ε-N-formyl-L-lysine were not substrates of trypsin. ε-N-Dimethyl-L-lysine derivatives did not inhibit the action of trypsin on a specific substrate. DL-Homolysine derivatives were hydrolyzed with kcat's one to two orders of magnitude lower than those of lysine derivatives, but their Km's were only 1.5–3 times higher. ε-N-Methyl-L-lysine derivatives were hydrolyzed at rates similar to those for DL-homolysine derivatives, and had Km's 25–115 times those of lysine derivatives. Plots of Km and kcat/Km versus side-chain length of the substrate for the ethyl esters of all the homologues of lysine and arginine indicated a correlation between these kinetic constants and side-chain length, and that the best substrate would have a side-chain length between those of lysine and arginine. Poly-ε-N-methyl-L-lysine was degraded to small peptides by trypsin.
Some properties of phosphodiesterase I from microsomes of rat intestinal mucosa
Authors: Hynie I. , Zbarsky S. H.
Published: December 2009

Some properties of phosphodiesterase I from microsomes of rat intestinal mucosa were studied using the p-nitrophenyl ester of thymidine 5′-phosphate as substrate. The optimum pH was found to be 9.5–9.6 in several buffer systems. The activity was destroyed at temperatures of 63–68°, depending on the pH of the medium. Mg2+, Ca2+, Sr2+, Ba2+, and Ni2+ had a variable activating effect while Al3+, Cu2+, and Be were inhibitory. The enzyme was inhibited completely by 10−5 M EDTA or EGTA in the assay system used. The evidence obtained suggested phosphodiesterase I is a metal-requiring enzyme. Glycine, bicine, and tricine inhibited the enzyme at higher concentrations. Cysteine, 2-mercaptoethanol, and dithiothreitol were strongly inhibitory at a concentration of 10−3 M. The Km of the enzyme was 8.5 × 10−4 M. In 6 M urea 40% of the activity of the enzyme was lost irreversibly after 24 h at 4°. In 3 M urea there was an immediate and reversible mixed competitive–noncompetitive inhibition and the Km was increased seven to eight times.
The reaction of 2,4,6-trinitrobenzene sulfonic acid with pancreatic elastase
Authors: Rao Leticia , Hofmann T.
Published: December 2009

The reaction of elastase with trinitrobenzene sulfonic acid was investigated in the pH range 9–12. Elastase was found to be inactivated by 2,4,6-trinitrobenzene sulfonic acid. The pH dependence of the pseudo first-order inactivation rate constant showed a pK of 10.3 and gave a Hill plot coefficient of 1.15. Trinitrophenol did not inactivate the enzyme. These results indicate that the inactivation is due to the covalent reaction of trinitrobenzene sulfonic acid with a single group in the enzyme. This group is not the N-terminal since the loss of N-terminal valine was considerably slower than the loss of activity at pH 10.5. The inactivation of elastase with 2,4-dinitrofluorobenzene also showed no correlation with the loss of the N-terminal. When the enzyme was exhaustively treated and fully inactivated with trinitrobenzene sulfonic acid at pH 10.5, the N-terminal valine and two out of three lysine residues were trinitrophenylated. No evidence for the loss of histidine was found. One of the tyrosine residues may be trinitrophenylated as judged from the molar extinction of the trinitrophenylated protein, but it has not been possible to isolate a trinitrophenylated tyrosine-containing peptide. The results can be interpreted in one of two ways: (a) trinitrophenylation of a group with a pK of 10.3, not involved in the activity, inactivates because the introduction of the trinitrophenyl residue causes a denaturation of the enzyme; or (b) a group with a pK of 10.3 controls the active conformation of the enzyme. The results do not exclude the possibility that the N-terminal plays an important role in the activity of the enzyme. Below pH 10.5 the reactivity of the N-terminal is low, indicating that it is buried.At pH 9.0 only the ε-amino group of lysine in position 224 reacted with trinitrobenzene sulfonic acid and full activity was retained. The second-order rate constant for the trinitrophenylation of this group was 25 times higher than that of the ε-amino group of the α-N-benzoyllysine.
Biosynthesis of Se-methylselenocysteine and S-methylcysteine in Astragalus bisulcatus
Authors: Chen D. M. , Nigam S. N. , McConnell W. B.
Published: December 2009

Biosynthesis of Se-methylselenocysteine and S-methylcysteine in Astragalus bisulcatus has been investigated using radiotracer methods. The results of feeding L-serine-U-I4C, DL-serine-1-14C, L-cysteine-U-14C, and L-methionine-methyl-14C to 2-month-old seedlings indicate that serine and cysteine are precursors of the three carbon chains, and that methionine methyl group is a precursor of the methyl group of the Se- and S-methylcysteines. It is suggested that in the biosynthesis of Se-methylselenocysteine from serine, Se-cysteine is formed first, and that Se-methylselenocysteine is formed from Se-cysteine by methylation. The incorporation of radioactivity into S-methylcysteine suggests a similar mechanism for its biosynthesis.
Phospholipids. IV. Studies on the antithromboplastic activity of phosphatidylserine preparations and their derivatives
Authors: Zitko V. , Okany A. , Palagy L.
Published: December 2009

Natural phosphatidylserine has antithromboplastic activity. This activity increases on storage, reaches a maximum, and then decreases. Evidence is presented which suggests that the change of the antithromboplastic activity is due to the autoxidation of phosphatidylserine. The increased activity can be further enhanced and stabilized by catalytic hydrogenation and decomposition of the residual peroxides of the autoxidized phosphatidylserine. A chemical oxidation of phosphatidylserine can be used instead of autoxidation.
Characterization of the esterases of canine serum
Authors: Ecobichon D. J.
Published: December 2009

Various techniques including electrophoresis, gel filtration, and titrimetric and spectrophotometric analysis with specific and nonspecific substrates and inhibitors were used to characterize the serum esterases of male and female dogs. Electrophoresis in starch gel yielded 10 distinct bands of activity: four butyrylcholinesterase bands, an esterolytically active albumin, and five bands of aliesterase and/or arylesterase activity. Gel filtration on Sephadex G-200 yielded two distinct peaks of activity, one containing the cholinesterase bands, and the second peak containing both arylesterase and aliesterase activity in close association with the serum albumin. Kinetic characteristics determined for the canine serum esterases included (1) optimal pH; (2) Km values for esters of choline, α-naphthol, and p-nitrophenol; and (3) average rates of hydrolysis of α-naphthyl acetate, butyrylcholine iodide, p-nitrophenyl acetate, and E600 by the sera of males and females. The sensitivity of the serum esterases to inhibition by various cations, and specific and nonspecific inhibitors was investigated. The organophosphate, E600, may be a highly specific substrate for the detection and quantification of arylesterase in a mixture of nonspecific carboxylesterases.
Interaction of δ-Chymotrypsin with Proflavine
Authors: Chen Wendy , Russo Salvatore F.
Published: December 2009

A 1:1 molar stoichiometry has been found for the noncovalent bonding of proflavine with δ-chymotrypsin. The dissociation constant at pH 7.6 is 0.025 mM which is similar to that found by Glazer for the reaction of proflavine with α-chymotrypsin. The difference spectrum produced by the proflavine–δ-chymotrypsin interaction has been studied as a function of pH. It would appear that a group with pK of approximately 6.5 is involved.
Metabolism of 15α-Hydroxyandrostenedione by the Perfused Human Placenta
Authors: Meeker C. I. , Stern M. D. , Solomon S.
Published: December 2009

14C-15α-hydroxyandrostenedione was perfused through two human placentas at the time of Caesarean section. The aromatization of 15α-hydroxyandrostenedione has been confirmed as a pathway for the placental production of 15α-hydroxyestrone and 15α-hydroxyestradiol in the human term placenta.
Partial Characterization and Cellular Localization of Two Deoxyribonucleases in the Small Intestine of the Rat
Authors: Liebbrman M. W. , Sullivan R. J. , Shull K. H. , Liang H. , Farber E.
Published: December 2009

We have partially characterized and localized two previously reported deoxyribonucleases from the rat small intestine. After separation of the crypt cells and muscle (the deep layer) from the villus cells (the superficial layer), the latter was found to contain a deoxyribonuclease I with a pH optimum around 6, and a molecular weight of 32 000 – 35 000. It was activated by Co2+, Mg2+, and Mn2+. Ligation of the pancreatic duct reduced the activity in intestinal extracts to about one-third of control levels. A deoxyribonuclease II with a pH optimum of 5.3–5.4 was found associated with the continuously dividing intestinal crypt cells. It was inhibited by Mg2+ and activated by EDTA. Ligation of the pancreatic duct was without effect on this enzyme. The deoxyribonuclease I is probably largely extracellular and serves a digestive function while the deoxyribonuclease II probably is related to intracellular DNA metabolism.
Resolution of Egg Yolk Lipoproteins by Chromatography on Thin Layers of Hydroxylapatite
Authors: Gornall D. A. , Kuksis A.
Published: December 2009

Egg yolk lipoproteins were separated into four major density classes by conventional ultracentrifugation, and each class was resolved further by thin-layer chromatography on hydroxylapatite. The plates were developed in special saturation chambers with phosphate buffers of 0.4, 0.6, 1.2, and 2.0 M for 2 h, and bands were located by exposure to iodine vapor or by spraying with ninhydrin. The low density fraction and the low density fraction of the granule each gave two subfractions, while the phosvitin–lipovitellin fraction yielded three components and the water-soluble fraction four components. The additional resolution apparently was due to differences in lipid and protein composition and structure, as well as to the content of protein-bound phosphorus. The described separations offer special advantages for the study of the lipid parts of the lipoprotein complexes.
The Use of Nucleotide-Sugar: Glycoprotein Glycosyltransferases to Assess Golgi Apparatus Function in Morris Hepatomas
Authors: Hudgin R. L. , Murray R. K. , Pinteric L. , Morris H. P. , Schachter H.
Published: December 2009

Enzymatic assays for CMP-sialic acid: glycoprotein sialyltransferase and UDP-N-acetylglucosamine: glycoprotein N-acetylglucosaminyltransferase were performed on crude homogenates of three Morris hepatomas (7777, 7800, and 5123D), and on liver homogenates from the host animals and normal Buffalo strain rats. It was found that sialyltransferase activities were greatly decreased in the most rapidly growing tumor (hepatoma 7777) and were decreased to a lesser extent in the more slowly growing hepatoma 7800; enzyme activities in hepatoma 5123D, another relatively slow growing tumor, were not significantly different from control values. Sialyltransferase activities were significantly elevated in the livers of all the tumor-bearing animals and were especially high in the livers of animals carrying hepatoma 7777; these elevations may be related to increased plasma glycoprotein synthesis by liver secondary to the inflammatory stimulus generated by the tumors. In contrast to the sialyltransferase analyses, N-acetylglucosaminyltransferase activities in tumor homogenates were very similar to control values for all three hepatomas. When the data are expressed as ratios of sialyltransferase activity to N-acetylglucosaminyltransferase activity, two of the three tumors show highly significant decreases of this ratio compared to either control or host livers. Since these glycosyltransferases have previously been shown to be located in the Golgi apparatus of normal rat liver where they function in the biosynthesis of glycoproteins, the above results have been interpreted to indicate a shift in the function of the Golgi apparatus in certain Morris hepatomas as compared to normal livers. Finally, glycosyltransferase assays and electron microscopy have been used to demonstrate the feasibility of preparing Golgi-enriched fractions from all three hepatomas by methods previously applied to normal rat liver.
Reversal of the Interference by Sucrose on Protein Determination by the Biuret Method
Authors: Arora D. J. S.
Published: December 2009

Sucrose interference is large when small quantities of proteins are determined by the biuret method. Precipitation with perchloric acid eliminates the effect of sucrose.
Studies on Porcine Haptoglobin and its Complex with Human Hemoglobin
Authors: Fraser Ian H. , Smith David B.
Published: December 2009

Haptoglobin, prepared from pig plasma, was separated into two components by gel filtration in guanidine hydrochloride solution, after reduction and aminoethylation. These were identified as homologous with the heavy and light polypeptides of human haptoglobin 1-1. Similar treatment of the complex formed by pig haptoglobin with human hemoglobin enabled separation of the hemoglobin and haptoglobin polypeptides.
Regulation of Lysosomal Enzymes. II. Reversible Adaptation of Intestinal Acid Hydrolases During Starvation and Realimentation
Authors: Desai I. D.
Published: December 2009

The nutritional role of intestinal lysosomal enzymes in the regulation of reversible adaptation to starvation and realimentation by various dietary treatments is investigated. When rats were starved for periods ranging from 72 to 120 h, the specific activity of the representative lysosomal hydrolases, viz. acid phosphatase, β-glucuronidase, β-galactosidase, arylsulfatase, and cathepsin D of small intestinal mucosa, progressively increased reaching levels two to four times higher than the fed controls. During the starvation period of only 72 h, more than 60% of the intestinal weight and about 75% of the mucosal protein was lost. On realimentation with a complete diet such as laboratory rat chow, the specific activity of the intestinal lysosomal enzymes rapidly returned to normal and the lost weight of the intestinal tissue and of the body as a whole was restored. The regulatory effect of major dietary nutrients such as carbohydrates, proteins, and fats on the starvation-induced breakdown of the intestinal tissue reserves is presented and discussed. The findings of this study indicate that changes in the specific activity of the intestinal lysosomal enzymes are associated with adaptive processes requiring rapid turnover of body tissues for maintenance and survival of the animal during starvation and/or during conditions when dietary supply of certain nutrients is limited.
Sex Hormone Control Mechanisms. II. Influence of Estrogen and Progesterone on the Activities of Key Enzymes Involved in the Carbohydrate Metabolism of Chicken (Gallus domesticus) Oviducts
Authors: Campbell L. D. , Yu J. Y.-L. , Stothers S. C. , Marquardt R. R.
Published: December 2009

Sex hormones were administered to chicks by repeated intramuscular injections, and the activities of certain enzymes of the oviduct were determined and compared with those of nontreated chicks and those of mature laying and nonlaying hens. The administration of estrogen in combination with progesterone resulted in enhanced activities of phosphofructokinase (ATP : D-fructose-6-phosphate 1-phosphotransferase, EC 2.7.1.11), aldolase (fructose-1,6-diphosphatase D-glyceraldehyde-3-phosphate-lyase, EC 4.1.2.13), and fructose-1,6-diphosphatase (D-fructose-1,6-diphospnate 1-phosphohydrolase, EC 3.1.3.11). Furthermore, estrogen treatment alone resulted in a lesser, but significant, response in phosphofructokinase and aldolase activities and no response in fructose-1,6-diphosphatase activity, while progesterone administered alone had no effect. The data indicate that in addition to enhancing the protein synthetic capacity of chick oviduct cells, estrogen or a combination of estrogen and progesterone also regulate, either directly or indirectly, the metabolic activity of these oviduct cells by increasing the activity of some enzymes involved in carbohydrate metabolism.
Lymphocyte and Platelet Adenylcyclase
Authors: McDonald John W. D.
Published: December 2009

Lysates of platelets and lymphocytes possess adenylcyclase activity which is stimulated by sodium fluoride. Platelet adenylcyclase activity is also stimulated by prostaglandin E-1. An apparent effect of prostaglandin on lymphocyte adenylcyclase was shown to be due to low-level platelet contamination of the lymphocyte preparation.
The Electron Paramagnetic Resonance Spectrum at 77 °K of Lyophilized and Frozen Solutions of Horse Heart Ferricytochrome c
Authors: Morton R. A. , Bohan T. L.
Published: December 2009

The electron paramagnetic resonance spectrum of a repurified, commercial preparation of horse heart ferricytochrome c was measured both in frozen solution, and as a lyophilized powder at 77 °K. The solution spectrum agreed with previous measurements reported at 20 °K. The lyophilized powder had both high-spin (g ~ 6), and low-spin components. The latter were significantly different from that of the solution spectrum. Thus, the molecular structure of the protein near the iron atom must be considerably, but reversibly, distorted by drying. In addition, it was suggested that a g = 4.29 line was due to acid-denatured forms of cytochrome c present as an impurity.
The Metabolism and Conjugation of 15α-Hydroxylated Phenolic Steroids by the Rabbit
Authors: Polakova A. , Gwilliam C. , Quamme G. A. , Williamson D. G. , Layne D. S.
Published: December 2009

The rabbit converts administered 15α-hydroxyestrone and 15α-hydroxyestradiol-17β to 15α-hydroxy-estradiol-17α. This conversion is incomplete when large amounts of the steroids are injected. All three of the 15α-hydroxylated estrogens are excreted as monoglucuronides, and no evidence was found for the formation of double conjugates or of N-acetylglucosaminides. The rabbit therefore differs from the human in that it does not transfer N-acetylglucosamine to the 15α-hydroxyl group of the phenolic steroids, and the presence of this group prevents the usual transfer of this sugar to the 17α-hydroxyl group.
Steroid Δ4-Reductases of Pig Liver: Partial Purification of Testosterone 5β-Reductase
Authors: Nduaguba J. C. , Clark A. F.
Published: December 2009

Intracellular distribution studies of steroid Δ4-reductase activity in female pig liver were done using testosterone as substrate. About 60% of the total enzyme activity was found in the 100 000 × g soluble fraction. Labelled 17β-hydroxy-5β-androstan-3-one but not 17β-hydroxy-5α-androstan-3-one was isolated from the incubation of 1,2-3H-testosterone with the 100 000 × g supernatant fraction, indicating the presence of 5β-reductase activity. 5β-Reduction may play an important role in the inactivation of some Δ4-3-ketosteroids in the pig liver.Evidence that 5β-reductases differing in substrate specificity are present in the soluble fraction includes (a) variation in the ratios of enzyme activities for several Δ4-3-ketosteroids in different (NH4)2SO4 fractions obtained from the 100 000 × g soluble fraction, (b) kinetic data showing that the maximum velocity for an equimolar mixture of testosterone and hydrocortisone is the sum of the maximum velocities for the substrates when used singly, and (c) separation of the enzyme activity specific for testosterone from that specific for hydrocortisone by use of Sephadex G-100 and hydroxylapatite chromatography.Utilizing (NH4)2SO4 precipitation and chromatography on Sephadex G-100 and hydroxylapatite, a 105-fold purification of testosterone Δ4-5β-reductase from the 100 000 × g supernatant fraction has been attained. The presence of 5 mM β-mercaptoethanolamine increased the stability of the enzyme.
Effect of Repeated Starvation on Serum Insulin Level and the Enzyme Overshoot in Liver
Authors: Szepesi B. , Berdanier C. D. , Diachenko S. K. , Moser P. B.
Published: December 2009

The responses of serum immunoreactive insulin (IRI), glucose-6-phosphate dehydrogenase (G6PD), malic enzyme (ME), and total liver lipid to starvation and refeeding were studied in male Wistar rats. All parameters studied were reduced by starvation. In rats refed a high-carbohydrate diet for 2 days, G6PD, ME, and total liver lipid rose to over twice the ad-libitum-fed values; IRI was approximately at or below ad-libitum-fed values. Treatment of rats with 8-azaguanine prevented the overshoot of G6PD, ME, and total liver fat, but did not prevent return to ad-libitum-fed values. A second cycle of starvation refeeding induced an even greater overshoot of G6PD and ME, while the overshoot of total liver lipid was not different from that observed after the first starve–refeed cycle, and serum insulin levels were again normal. Treatment with 8-azaguanine during the second refeeding only allowed G6PD and ME to reach approximately twice ad-libitum-fed values. Two basic conclusions were drawn from the data: (1) that hyperlipogenesis in starved–refed rats is not due to continuous hyperinsulism, and (2) that the newly formed RNA responsible in part for the enzyme overshoot is relatively stable.
Isolation of a Basic Protein Antigen of Low Ragweed Pollen
Authors: Griffiths B. W. , Brunet R.
Published: December 2009

A fractionation method has been developed for the preparation of a basic protein antigen (BPA-R) from ragweed pollen. This antigen possesses potent allergenic activity. It is homogeneous by gel filtration on Sephadex G-100 and carboxymethyl-Sephadex chromatography. By the former technique the molecular weight of the BPA-R was estimated to be 28 000. Judged by its physicochemical and immunologic properties the BPA-R appears to be distinct from several other highly purified antigens currently isolated from ragweed pollen.The BPA-R and AgE (a highly purified acidic protein antigen) when tested together in Ouchterlony analysis displayed reactions of typical partial immunological identity with rabbit anti-ragweed sera which were produced by hyper immunization utilizing complete Freund's adjuvant. In contrast rabbit antisera, produced on a shorter schedule with BPA-R (using incomplete Freund's adjuvant), failed to react by Ouchterlony test with the AgE but reacted strongly with the BPA-R. Further immunological as well as chemical studies will be necessary to elucidate the antigenic relationship between these moieties.
Optical Rotatory Dispersion and Circular Dichroism Studies on Ocular Lens Proteins
Authors: Jones Howard A. , Lerman Sidney
Published: December 2009

A study has been made of the circular birefringence and circular dichroism of alpha and gamma crystallins obtained from dogfish lenses. The Cotton effects in the optical rotatory dispersion and circular dichroism spectra are correlated with maxima in the electronic absorption spectra. On the basis largely of this correlation, the transitions which give rise to the Cotton effects have been assigned to various chromophores. It is concluded that the secondary structure of these proteins is made up of beta structure and the unordered conformation. Little or no α-helical conformation seems to be present.
The Sequence Distribution of O2′-Methylribose in Yeast Ribosomal RNA
Authors: Lau R. Y. , Lane B. G.
Published: December 2009

(1) There are 16 possible alkali-stable Nm–N sequences in which N is any of the four major ribonucleosides and Nm is the O2′ -methyl derivative of any of the four major ribonucleosides. All of these sequences have been found to be present in both 17 S and 26 S RNA from exponentially dividing yeast cells.(2) The relative amounts of the different Nm–N sequences in 17 S RNA differ from the corresponding proportions found for 26 S RNA.(3) Some of the Nm–N sequences in 17 S RNA are not present in every polynucleotide chain and possible interpretations of this finding are discussed.(4) By comparing the present results of analyses of plant RNA with earlier analyses of animal and bacterial RNA, it is possible to perceive underlying regularities for the patterns of O2′-methylation in animal, plant, and bacterial RNA.
Effects of X-Irradiation on the Various Enzymic Functions of NADP+-Linked Isocitrate Dehydrogenase
Authors: Holland Peggy , Little Clive
Published: December 2009

Highly purified NADP+-linked isocitrate dehydrogenase was X-irradiated in dilute aqueous solution and proved to be very radiosensitive, being inactivated with a G value of 0.5. The catalytic functions of the enzyme were destroyed at the same rate. However, unlike most allosteric enzymes, the allosteric properties were not more radiosensitive than the catalytic properties. X-ray inactivation was associated with the destruction of seven to eight sulfhydryl groups. The first four sulfhydryls destroyed were extremely radiosensitive, having G values for destruction of almost 2. Comparison with several reagents known to attack sulfhydryl groups suggested that sulfhydryl destruction was sufficient to account for the X-ray response of the enzyme. Destruction of the single essential methionine did not seem significant in the inactivation. X-ray inactivation was also associated with increases in the Km values for isocitrate and manganese.
Some Enzymatic Properties of Trypsin after Reaction with 1-Dimethylaminonaphthalene-5-sulfonyl Chloride
Authors: Franklin James G. , Leslie James
Published: December 2009

At pH 7–8, and at 0–25°, 1-dimethylaminonaphthalene-5-sulfonyl chloride reacts with an isoleucine and a lysine residue, with some additional reaction at a tyrosine residue, in trypsin. The enzymatic activity of the modified trypsin toward N-benzoyl-DL-arginme p-nitroanilide is greater than that of the unmodified enzyme, but there is little or no change in activity toward N-benzoyl-L-arginine ethyl ester, N-benzoyl-DL-arginine ethyl ester, and N-benzoyl-L-arginine amide. The enhanced activity does not appear to be due to a general increase in the catalytic part of the mechanism of trypsin action, but rather to an increased binding of N-benzoyl-L-argimne p-nitroanilide to the enzyme.
Isolation of Dinitrophenyl-Specific Antibodies from Bovine Colostrum
Authors: Froese A.
Published: December 2009

Antibodies specific to the 2,4-dinitrophenyl determinant were elicited in the colostrum of a cow, by immunization with a conjugate of porcine gamma globulin and 2,4-dinitrobenzene sulfonate. These antibodies were found to be associated with the IgG and IgM classes of colostral whey and could be specifically isolated, using a cellulose immunosorbent. The binding of the dye-hapten 1-hydroxy-4-(2,4-dinitrophenylazo)-2,5-naphthalene disulfonate by the nonprecipitating IgG antibodies yielded an equilibrium constant of 7.5 × 104 M−1 and revealed that the antibody molecule had two combining sites.
A Spin Label Study of the Influence of Cholesterol on Phospholipid Multibilayer Structures
Authors: Hsia Jen-Chang , Schneider Henry , Smith Ian C. P.
Published: December 2009

Cholesterol increases the degree of order indicated by the spin-labelled derivative of cholestane (3-spiro-(2′-(N-oxyl-4′,4′-dimethyloxazoIidine))-cholestane) in a hydrated multibilayer structure of egg lecithin. Estimates of the angle between the long axis of the label and the lamellar plane were obtained using a simple model for the effect of this angle on the angular dependence of the hyperfine splittings of the spin label. The deviation from perpendicularity was ~28 + 5° in the absence of cholesterol. This deviation decreased to a limiting value of ~10 ± 3° at 25 mole% cholesterol. The limiting deviation was smaller when dipalmitoyl lecithin was substituted for egg lecithin, indicating the importance of the nature of the acyl residues in this effect.The rotational frequency of the spin label about its long axis depended upon whether or not the multibilayer structure had been in contact with an aqueous phase. Before wetting, this frequency was « 75 × 106 s−1; after wetting it was » 75 × 106 s−1.
Multiple Molecular Forms of Avian Aldolases. VI. Enzymatic Properties and Amino Acid Composition of Chicken Liver Aldolase and Comparative Immunochemical Properties
Authors: Marquardt Ronald R.
Published: December 2009

Several properties of pure chicken (Gallus domesticus) liver aldolase (fructose-1,6-diphosphate D-glycer-aldehyde-3-phosphate-lyase, EC 4.1.2.13) were determined. The enzyme shows a broad pH optimum peaking around pH 7.7, is stable over a wide pH range, and has a temperature coefficient Q10 of 2.3. The enzyme is stable at 48 °C for 10 min and is almost completely inactivated at 55 °C.The amino acid composition of pure chicken liver aldolase was determined. Its composition was compared with those of chicken muscle and chicken brain aldolases and with the corresponding rabbit aldolase enzymes.Antiserum was prepared against purified chicken liver aldolase and the immunological properties of aldolases from various tissues and species were examined. In the chicken, the antiserum has a very pronounced reaction with liver and kidney aldolases but only a weak cross-reaction with either the brain or the muscle enzyme. The antiserum readily cross-reacts with other avian liver aldolases (turkey and budgerigar) but not with mammalian liver aldolases (rat and rabbit).
The Effect of Ligand Binding on the Optical Rotatory Dispersion of Lactoperoxidase
Authors: Maguire R. James , Dunford H. Brian
Published: December 2009

The optical rotatory dispersion of lactoperoxidase and its cyanide and fluoride complexes was studied over the spectral region 210–500 nm, and that of the azide complex from 350–450 nm. Results of the measurements of the reduced mean rotation at 233 nm lead to the conclusion that there are no significant changes in gross protein conformation upon the binding of these ligands to lactoperoxidase. Lactoperoxidase was estimated to have 17% α-helical character at pH 7.0. Results of studies in the Soret region indicate that lactoperoxidase, unlike horseradish peroxidase, hemoglobin, and myoglobin, exhibits a negative Cotton effect as do its cyanide, azide, and fluoride complexes. The binding of these ligands apparently causes an alteration of the geometry of the heme group with respect to the protein moiety.