High Protein Fraction Research Articles

The chemical form of cadmium in subcellular fractions following cadmium exposure.

Wistar rats were given drinking water containing 250 ppm Cd for 12 months. After excision of the kidney and liver, the organs were subfractionated into nuclear, mitochondrial, microsomal and cytosol fractions, and the chemical forms of Cd in the subcellular fractions were examined. Although approximately 90% of the total Cd was present in the cytosol, in the form of metallothionein, 3-5% was also present in the mitochondrial fraction and 5-7% in the microsomal fraction from both organs. By Sephadex G-75 gel filtration, after solubilizing the particulate fractions with sodium deoxycholate, approximately 89% of Cd in the microsomal fraction and 94% in the mitochondrial fraction eluted with the same retention time as that of metallothionein in both liver and kidney, while the remainder was found in a high molecular weight protein fraction. The Cd that eluted with the high molecular protein fraction may be involved in dysfunction in subcellular organelles. For estimating the toxicity of Cd associated with the high molecular weight fraction, rat liver microsomes and CdCl2 were mixed with 1% of sodium deoxycholate and the protein-Cd complex produced was isolated by eluting with Sephadex G-75. This complex had a strong toxicity toward the alcohol dehydrogenase activity (SH-enzyme), and the Ki values of the Cd-protein complexes decreased with increased amount of Cd bound to the microsomal protein fraction. The above results suggest that loosely bound Cd increased in the case of higher Cd/protein and plays a toxic role in the living cells.

Air Classification and Extrusion of Navy Bean Fractions

ABSTRACTPilot plant air classification or raw beans (Phaseolus vulgaris) produced high‐protein fractions (HPF) containing 51.4% protein and high‐starch fractions (HSF) having about 50% starch. Particle size distribution analyses and scanning electron micrographs confinned the fractionation of starchy and proteinaceous materials. Extruded corn/HSF blends had lower water absorption and water solubility indices, higher protein content, and a better balanced amino acid pattern than pure corn extrudates. Extrusion‐texturized vegetable proteins obtained by substituting HPF for defatted soy flour at levels of 10, 20 and 30% had similar functional properties as the 100% texturized soy. Air classification followed by extrusion‐cooking is a feasible alternative for dry‐processing of beans into products for human consumption.

Physicochemical Characteristics of Dry‐Roasted Navy Bean Flour Fractions

ABSTRACTNavy beans (Phaseolus vulgaris) were dry roasted in a particle‐to‐particle heat exchanger, dehulled by air aspiration, pin‐milled and air‐classified to yield whole, hull, high protein, and high starch flour fractions. Proximate analysis, color, ENDF, starch, nitrogen solubility indices, and oligosaccharide contents of these flour fractions were determined. Compositional differences were demonstrated among all flour fractions. Stachyose was the major oligosaccharide in all fractions and was highest in the protein fraction. Substituting 10% bean flours for wheat changed water absorption and dough stability for all fractions. Dry roasted navy bean flour fractions appeared to be suitable ingredients for use in appropriate food systems.

Characteristics and utilization of dry roasted air‐classified navy bean protein fraction

AbstractNavy beans,Phaseolus vulgaris, were dry roasted in a particle‐to‐particle heat exchanger, dehulled by air aspiration, pin‐milled and air‐classified to yield a high protein fraction. Proximate analyses, nitrogen solubility indices and oligosaccharide contents of this high protein fraction as influenced by processing parameters which affected final product temperature were determined. Farinograms of wheat/bean protein fraction composite flours were run. A high‐protein bean flour fraction was selected from these dry and roasted treatments and used in product development. Quality characteristics and consumer acceptability of high‐protein prototype products were evaluated. Results of this research indicate that the dry roasting process influences the characteristics of the air‐classified protein fraction. Flour color, nitrogen solubility and dough mixing properties were most greatly influenced by roasting time and temperature. Increased roasting resulted in increased browning and decreased nitrogen solubility and dough mixing stability. Wheat flour bread products, substituted with low levels of high‐protein bean flour, were of high quality.

Effects of proteolytic enzymes on steroid release from rat adrenal zona glomerulosa tissue: Evidence for novel steroid-protein complexes

Trypsin (2mg/ml) added to conventional incubations of rat adrenal capsules (largely glomerulosa) reproducibly increases the amount of free extractable aldosterone (aldo) and 18-hydroxycorticosterone (18-OH-B) in incubation media, but has no effect on capsule cell suspensions formed by collagenase incubation. The effect is abolished by the addition of a trypsin inhibitor, but is still seen in the absence of de novo steroidogenesis. Qualitatively similar results were obtained with capsule homogenates and high speed supernatant fractions, and chromatography of the high speed supernatant protein fraction on Sephadex G-50 gave a number of minor fractions and one major fraction which yielded free aldo on incubation with trypsin. The results indicate the existence of storage forms of aldo and 18-OH-B which are extremely tightly bound to protein. Such steroid-protein complexes appear to be of an entirely novel kind, and are quite distinct from the familiar receptor type complexes. The findings support previously proposed mechanisms for aldo synthesis and secretion.

Plasma retinol and retinol binding protein concentrations in patients on maintenance haemodialysis with and without vitamin A supplements.

Plasma retinol and retinol-binding protein (RBP) concentrations have been estimated in patients on maintenance haemodialysis over a 4-year period. For the first 2 years multivitamin supplements containing vitamin A were taken, and for the second 2 years no vitamin A supplements were given. Mean plasma retinol concentrations decreased significantly but only from 3.8 times normal to 3.1 times normal after vitamin A supplements stopped. There was no significant change in th high plasma RBP levels. Ultracentrifugation of plasma at a salt density of 1.21 showed that nearly all the retinol was associated with RBP in the high density protein fraction, as it is with normal subjects. Column chromatography confirmed that there was no increase in plasma retinyl esters in the renal failure patients, as is found in hypervitaminosis A due to drug overdosage. The high plasma retinol and RBP levels remained remarkably stable in individual patients throughout the 4-year study. The increase in plasma RBP was possibly related to residual urine output. The results are compatible with a feedback mechanism whereby the extent of the increase in plasma RBP as renal failure develops controls the consequent high plasma level of retinol.

Effect of detoxification treatments on the proteins of guar meal.

Guar meal was subjected to various detoxification pretreatments, and their effect on the proteins was determined by chemical and physicochemical methods. Ethanol treatment removed the high and low molecular weight protein fractions and decreased the carbohydrate content. These two protein fractions were also not observed in the proteins of the autoclaved samples, while there was no appreciable change in the carbohydrate content. Autoclaving also reduced the available lysine content of the total meal protein. Acid treatment dissociated the high molecular weight protein fractions and reduced both the available lysine and carbohydrate contents.

The chemical form of cadmium in microsomal and mitochondrial fractions from rat liver and kidney after long term administration of cadmium chloride

Wistar rats were given drinking water containing 250 ppm cadmium (Cd) for 12 months. After excising the kidney and liver, the organs were subfractionated into nuclear, mitochondrial, microsomal and cytosol fractions, and the chemical forms of Cd in the subcellular fractions were examined. Although approx. 90% of the total Cd was present in the cytosol, in the form of metallothionein, 3–5% was also present in the mitochondrial fraction and 5–7% in the microsomal fraction from both organs. Each membrane fraction was washed 3 times and there was no contamination of metallothionein from the cytosol according to Cd/protein ratios. By Sephadex G-75 gel filtration, after solubilizing the particulate fractions with sodium deoxycholate, approx. 89% of Cd in the microsomal fraction and 94% in the mitochondrial fraction eluted with the same retention time as that of metallothionein in both liver and kidney, while the remainder was found in a high molecular weight protein fraction. The Cd eluted with the high molecular protein fraction might be involved in dysfunctions in subcellular organelles.

Binding of juvenile hormone, methoprene and hydroprene to haemolymph proteins of larvae of the southwestern corn borer, Diatraea grandiosella

The binding of tritiated juvenile hormone I, and the juvenile hormone mimics, methoprene and hydroprene (alkyl-3,7,11-trimethyl-2,4-dodecadienoates), to proteins of the haemolymph of larvae of the southwestern corn borer, Diatraea grandiosella, was examined in vitro. The techniques of analytical isoelectric focusing, disc electrophoresis and gel filtration were employed. Juvenile hormone was shown to bind to a high affinity low molecular weight protein in the haemolymph of diapausing larvae. This protein has an apparent dissociation constant of about 3.1 × 10 −7 M at 4°C, an isoelectric point of about 4.85, and a molecular weight of about 3 × 10 4 . Juvenile hormone was also bound to at least one low affinity high molecular weight protein fraction. High juvenile hormone binding activity was found in the haemolymph of newly diapaused larvae. Methoprene and hydroprene bound selectively to a different protein fraction present in the haemolymph of pre-diapausing larvae. This protein fraction had a low relative mobility when subjected to electrophoresis at pH 8.9. A preliminary study of the binding of tritiated juvenile hormone I to proteins of the haemolymph of pre-diapausing larvae of the southern cornstalk borer, Diatraea crambidoides, was also conducted in vitro. A low molecular weight binding protein with identical electrophoretic properties to that of D. grandiosella was present. The high molecular weight binding protein fraction of the two species, however, had different electrophoretic properties.

A cyclic AMP binding-protein from barley seedlings

A protein fraction extracted from barley seedlings was shown to bind 3′:5′-cyclic AMP. The binding effect is real and not due to interference with the standard binding-protein assay used. Evidence is presented that this is a specific binding-protein; even at high concentrations other protein fractions from the same source showed no affinity for cyclic AMP. None of a range of cyclic and non-cyclic nucleotides that were examined exhibited a degree of binding with the protein comparable to that with cyclic AMP. The cyclic AMP/binding protein complex has a K d of 8 nM. This complex eluted at an identical position in the elution sequence from a Sephadex G-150 column as the uncomplexed binding-protein. The barley binding-protein is in a fraction which also exhibits the enzymic activities of glucose 6-phosphatase, ATPase, 5′-nucleotidase, and fructose 1,6-diphosphatase.

Selection of an adequate protein-carbohydrate ratio by the domestic chick

Male broiler chicks, given access to high protein and high carbohydrate diet fractions, demonstrated an ability to select a protein-carbohydrate ratio sufficient to maintain growth at near control levels. The level of protein selected was below that present in either control diet and declined with age. It is hypothesized that a feedback loop involving learning controls the selection mechanism although the specific cues involved were not isolated in this study.

Über Lipoxygenase und Linolsäurehydroperoxid-abbauende Enzyme in Roggen

An enzyme fraction from rye containing lipoxygenase activity was investigated. The molecular weight of lipoxygenase was found to be about 102000. Two bands groups with isoelectric points between 5.1-5.5 and 5.8-6.4 were obtained by isoelectric focusing. Three isoenzymes could be separated by ion exchange chromatography. Lipoxygenase has optimum activity at pH 7.3-7.5 and predominantly forms 13-hydroperoxy-9-cis, 11-trans-octadecadienoic acid (13-LHPO). In rye the 13-LHPO is converted to alpha-ketols by a high molecular protein fraction. This isomerase converts the LHPO formed by rye lipoxygenase predominantly to 12,13-ketohydroxy acids. The Michaelis Constant of isomerase is 3-5 X 10(-5), using LHPO as substrate. At low protein concentrations the reaction velocity of LHPO-conversion increases linearly with protein concentration.

ENRICHMENT OF PASTA WITH COTTAGE CHEESE WHEY PROTEINS

ABSTRACTAddition of high protein fractions derived from whey to pasta to bring the protein content to 20% increases the PER of the pasta (macaroni) to that of casein. The flavor or texture quality of cooked enriched pasta is acceptable, and cheese or tomato sauce flavored enriched pastas are indistinguishable from controls. To avoid problems that would force major modifications in pasta processing and control equipment, it is recommended that the high protein fraction be insolubilized (denatured). The amino acids from these proteins are not changed by heat coagulation, pasta production, or cooking to a palatable table dish.

EFFECT OF EXOGENOUS LIPIDS ON ACTIVITIES OF THE RAT BRAIN CHOLESTEROL ESTER HYDROLASE LOCALIZED IN THE MYELIN SHEATH1

Abstract— Activity of cholesterol ester hydrolase localized almost exclusively in the myelin sheath (Eto & Suzuki, 1973a) was greatly affected by exogenous lipids added to the assay mixture. With isolated myelin as the enzyme source, phosphatidylserine was most effective in stimulating the activity. Other phospholipids were less effective. Efhanolamine phospholipid was slightly inhibitory and lysolecithin was strongly inhibitory. Differences in the fatty acid composition did not appear to account for such different effects. Glucosylceramide, galactosylceramide and digalactosylceramide were stimulatory while sulfatide, ganglioside and its asialo‐derivative were inhibitory. Saturated fatty acids were generally stimulatory while corresponding unsaturated acids were strongly inhibitory. In order for exogenous lipids to be effective they had to be added to the assay mixture as free dispersion. When heat‐inactivated myelin was used as the lipid source, no effect was observed, while equivalent amounts of a whole white matter lipid mixture was effective. Although phosphatidylserine was the most effective activator among the lipids tested, it could not completely replace sodium taurocholate present in the standard assay system. When isolated myelin was stored frozen, the activity of the enzyme declined gradually in the standard system without additional lipids. The stimulating effect of phosphatidylserine was greater for such partially inactivated enzyme sources, although it did not completely restore the activity to that of fresh preparations. When myelin was fractionated into basic protein, proteolipid protein and the high molecular weight acidic protein (Wolfgram) fractions, the last fraction contained most of the recovered activity. However, Wolfgram protein was less active than the intact myelin when assayed without additional lipid. The addition of phosphatidylserine completely restored the activity of this partially delipidated preparation.

Nucleic acid helix-coil transitions mediated by helix-unwinding proteins from calf thymus.

We have studied nucleic acid double helix destabilization mediated by purified calf helix-unwinding proteins, measuring ultraviolet hyperchromicity to detect helix melting. Both calf unwinding protein 1 (UP1) and a high salt eluting protein fraction are found to depress strongly the helix melting temperature (Tm) of the synthetic alternating copolymers poly[d(AT)] and poly[r(AU)], indicating that both DNA and RNA are recognized by these proteins. UP1 also destabilizes natural, GC-containing DNA helices, but to a smaller extent than observed with the above polymers. A simple model is presented to aid in the qualitative interpretation of the data, outlining the expected effect on the helix-coil transition of a protein ligand with differential affinity for the helix or coil form of nucleic acid. The observed helix-destabilizing effect of UP1 is dependent on the protein to nucleic acid ratio in an expected manner. Competition studies demonstrate a low, but appreciable affinity of UP1 for native DNA, opening the possibility that protein-mediated denaturation might be initiated by protein binding to the double helix. "Hairpin" helical regions of denatured DNA are strongly destabilized by UP1. Despite the fact that removal of these hairpin helices might greatly facilitate DNA renaturation, we failed to observe renaturation from the UP1-DNA complex after a switch to helix-stabilizing conditions. Thus, UP1 shows an important difference from its presumed prokaryotic analogue, T4 gene 32-protein. Possible in vivo functions of the calf proteins are discussed in light of these observations.

A BIOCHEMICAL MARKER IN A DOMINANT MOUSE TRAIT

Mice, heterozygous (Nn) for the Naked (N) trait, show patchy depilation, as a result of excessive breakage of the hair shafts. Hair from four Nn and from four normal mice belonging to the same strain, inbred for 57 generations with forced heterozygosis, was subjected to hydrolysis and amino acid analysis in triplicate. The mutant hair contained 20% less glycine and 27% less tyrosine than normal hair (p<0.001) as shown in the table, where the results are expressed in moles per cent.Whole hair was next extracted by reduction and alkylation. The Nn proteins thus extracted contained 30% less glycine and 40% less tyrosine than the normal proteins. These results suggest that Nn hair is deficient in the high glycine, high tyrosine protein fraction which is extracted by the procedures used and which comprises about 25% of the total protein in mouse hair. This mutation provides a rare biochemical marker in a dominantly inherited malformation.

Comparative Fractionation Studies of Cartilage Proteinpolysaccharides

Abstract Proteinpolysaccharides from bovine embryonic rudiments and calf nasal septa were compared by conventional ultracentrifugation in isotonic salt solutions, equilibrium density gradient centrifugation, and free flow electrophoresis. These methods are all suitable for use with large amounts of material and give reliable materials balances. Fractions obtained by any one method could be isolated and then refractionated by the other methods. The main distinction between fractions was in their composition. All fractions of the rudiment proteinpolysaccharide contained more protein than the corresponding nasal septum fractions. In gradient centrifugation, where the distribution of the proteinpolysaccharide was most influenced by protein content, much more of the rudiment proteinpolysaccharide than of the nasal septum proteinpolysaccharide was found in the less dense section of the gradient. Moreover, during conventional ultracentrifugation in isotonic salt solution, much more of the rudiment proteinpolysaccharide separated from the solution as a gel phase. All proteinpolysaccharide fractions obtained by the methods used in this study, regardless of composition, migrate to a characteristic peak area on electrophoresis. The protein-rich fractions of proteinpolysaccharide could be localized in the upper part of the gradient on centrifugation, in the separated, gel phase on conventional ultracentrifugation, and in the more slowly migrating section of the characteristic peak area on electrophoresis. Collagen is found in all high protein fractions and even, to a lesser extent, in the dense fraction of the rudiment proteinpolysaccharide. However, the collagen is not responsible for the distribution pattern of the proteinpolysaccharides on fractionation since that distribution is not altered by the removal of collagen with collagenase. The major component of protein-rich fractions from either tissue was shown to be proteinpolysaccharide by both analytical and electrophoretic criteria. The remixing experiments, which have attributed a specific linking function to a glycoprotein for the aggregation of proteinpolysaccharide, implicitly depend on the absence of proteinpolysaccharide in the low density fractions. Hence, the isolation of proteinpolysaccharide from these fractions questions the validity of such experiments as an unequivocal demonstration of a definitive role for the glycoprotein as a linking agent.

真菌の生化学的免疫化学的研究 (第9報)

We examined for the productive conditions of the extracellular antigenic glycopeptide which was excreted by Aspergillus fumigatus in culture medium and obtained the following results.Powerful antigenicity of culture filtrate of Asp. fumigatus growing on the sabouraud medium, yeast extract medium and the other synthetic media was observed after 10 days incubation especially, and it showed with appearance of a glycopeptide on the paperelectrophoretic pattern of 80% MeOH fraction of the filtrate after 3 or 4 days incubation.Total carbohydrate of the 80% MeOH fractions increased with development of culture and their sugar components were composed of galactose and mannose.From the results of the precipitin activity of the fraction eluted with water on Sephadex G-50 column, it was found that polysaccharide fraction showed high activity but protein fraction was very weak.

Protein Metabolism of Tissue Cells in vitro

Summary.1. In a treatment of the problems concerning the growth substances acting upon animal tissue cells in vitro it is pointed out that it is necessary to distinguish between the nature and mode of action of the growth promoting substances, termed embryonin, which are high molecular protein fractions present in an embryo extract, and the accessory growth substances of low‐molecular nature present in a variety of biological systems. Both of these groups were found to be essential to the growth of tissue cells in vitro.2. When studying the properties of the low‐molecular substances several difficulties are encountered which are inherent in the test method available and therefore unavoidable. The meaning of the term “accessory growth substances” for tissue cultures in vitro is discussed.3. The purification of the barley malt extracts previously investigated was carried further with only partial success. The activity of the fractions diminish as the purification proceeds. In some cases completely inactive fractions result, which when mixed regain a considerable activity indicating the presence of at least two groups of accessory growth substances essential to the tissue cells.4. From embryonic extracts it is possible to isolate as a stable dry powder purified preparation which is superior to the best malt extracts in producing normal growth and appearance of the tissue cultures.