Low Molecular Weight Protein Fraction Research Articles

Chemical characterization of peach kernel oil and protein: Functional properties, in vitro digestibility and amino acids profile of the flour

Kernels of peach fruits contain 54·5% and 27·5% oil and protein, respectively, but ash and total carbohydrates were quite low. Total lipids contained 98% triglycerides; sterols and polar lipids constituted 0·41% and 1·1% respectively. Fatty acid composition revealed the presence of oleic (63·8%) and linoleic (15·4%); there was 20·7% of saturated fatty acids. The defatted flour showed good fat absorption and emulsification properties but water absorption and foaming properties were slightly low. The solubility of protein was high at both acidic and alkaline pH with a minimum solubility at pH 4·3. Albumins were the major protein fractions (60%) followed by non-protein nitrogen (17%); globulins and glutelins constituted 9·5% and 8·2%, respectively, and prolamines only 6·0%. The protein was highly digested by pepsin-pancreatin followed by pancreatin and pepsin and the lowest digestibility was by trypsin. The protein is rich in lysine, leucine, isoleucine, valine, threonine, basic and acidic amino acids but poor in methionine. PAGE showed five bands with different relative mobilities and they contained both high and low molecular weight protein fractions.

Isolation and characterization of low-molecular-weight protein from safflower seed (Carthamus tinctorius L.)

The low molecular weight (LMW) protein fraction, often termed the 2s component, was isolated from safflower seed (Carthamus tinctorius L.). The LMW protein fraction, even though it sediments as a single peak with an S20,vwa lue of 1.5 S in the analytical ultracentrifuge, appears to be a group of several LMW proteins as indicated by polyacrylamide gel electrophoresis and DEAE-cellulose chromatography. The protein was characterized in terms of ultraviolet and fluorescence spectra, extinction coefficient, sedimentation coefficient, carbohydrate and phosphorus contents, several enzyme activities, and secondary structure. The protein has nearly 30% a-helix and 47% @-sheet in its secondary structure. Amino acid analysis of the protein indicates that the protein is rich in glutamic acid or glutamine and arginine. A chromophore is present along with the protein. The optical properties of the protein change upon dialysis due to dissociation of the chromophore.

Chemical and nutritional changes in soybean during germination

Germination of soybeans for 6 days depressed the trypsin inhibitor activity by 32%. Oil content, acid value, iodine value and total unsaturated fatty acids of soybean oil decreased while peroxide value and total saturated fatty acids pronouncedly increased. Percentage of palmitic and linoleic fatty acids were highly affected during germination. Analysis of the hydrocarbon and sterol classes revealed n-tricosane and β-sitosterol to be, respectively, the major components. Marked changes in both hydrocarbon and sterol constituents occurred upon germination. Different periods of soybean germination increased non-protein nitrogen and decreased reducing sugars. However, total ash content decreased due to soaking; thereafter it was constant for the rest of the germination period. Different amino acid components increased due to germination when compared with their content in dry seeds. The protein solubility of germinated soybean was higher in H 2 O, 5% NaCl and 0·02 m NaOH than it was in ungerminated seeds. Water and oil absorption, emulsification and foaming capacities of soybean were increased after germination. A marked improvement in digestibility by pepsin was noticed while tryptic digestibility was still low (38%). PAGE patterns showed drastic changes in both low and high molecular weight protein fractions due to germination.

Proteinuria in the dog: a clinicopathological study in 51 proteinuric dogs

In 51 dogs with predominantly massive urinary protein loss, the daily loss was quantified and glomerular and tubulointerstitial lesions from renal biopsies were characterised and graded using histology, immune fluorescence and electron microscopy. The highest median daily urinary protein loss occurred in dogs with membranous glomerulonephritis (median 380 mg kg-1 d-1) and renal amyloidosis (median 257 mg kg-1 d-1). Although in nine febrile dogs the urinary protein loss was transient, both glomerular and tubular lesions were diagnosed in five and seven of these dogs, respectively. The pattern of urinary proteins was determined using sodium dodecyl-sulphate polyacrylamide gel electrophoresis. The albumin fractional clearance (FC) was raised in 46 dogs, whereas the FCS of the low molecular weight (MW) protein fraction (MW less than 66,000) and high molecular weight protein fraction (MW more than 66,000) were raised in 42 and 28 dogs, respectively. Both the high molecular weight protein FC and albumin FC significantly correlated to the grade of glomerular lesions, whereas the low molecular weight protein FC only moderately significantly correlated to the grade of tubular lesions. The selectivity index, calculated as (formula; see text) did not differentiate between the various forms of glomerulopathies. The urinary lysozyme concentration was significantly correlated to the grade of tubular lesions. It is concluded that although quantitative and qualitative measurements of urinary proteins can provide additional clinical information, they do not have a reliable predictive value and histopathological examination of renal tissue is still necessary for the final diagnosis.

Copper-binding proteins in liver of bluegills exposed to increased soluble copper under field and laboratory conditions.

Livers from bluegills exposed to increased soluble copper (Cu) under field and laboratory conditions were analyzed to determine the concentration and distribution of Cu in metalloproteins of different molecular size. Analyses were performed on bluegills collected from the impoundment of the H. B. Robinson Steam Electric Plant (Florence, SC) near the effluent discharge from the power plant, near the water intake to the cooling system, and from a control pond as well as on bluegills exposed under controlled laboratory conditions. Metalloproteins were separated into low molecular weight (LMW), intermediate molecular weight (IMW), and high molecular weight (HMW) fractions by using high-performance liquid chromatography. In the field-exposed bluegills, Cu concentrations in the LMW, IMW, and HMW fractions were highest in bluegills from the discharge site and lowest in those from the control pond. In the laboratory-exposed bluegills, Cu concentrations in the fractions increased with exposure concentration and time. Concentrations of Cu in the LMW protein fraction and pellet of bluegills exposed to 160 micrograms Cu/L appeared to plateau with long exposure times, whereas those in the HMW fraction continued to increase. Bluegills maintained in 80 micrograms Cu/L water at pH 5.5 accumulated lower concentrations of Cu in the LMW and pellet fractions and higher amounts in the HMW than in those maintained in 80 micrograms Cu/L at pH 7.0. Mortality was dependent on exposure concentration and duration and was higher in bluegills maintained in water at pH 5.5 than at pH 7.0.

Electrical stimulation—Effects on the protein in the ventral nerve cord of cockroach,Periplaneta americana

Changes in protein concentration due to electrical stimulation have been investigated at different time intervals in cockroach ventral nerve cord. Significant increase in total protein concentration was observed at 15 min interval of stimulation. Increase in protein concentration was also observed in ventral nerve cords incubated with exogenous glucose. Microdisc Polyacrylamide gel electrophoresis at 15 min interval has revealed an increase in low molecular weight protein fractions in nerve cords from stimulated group. The data clearly depict the changes in protein due to electrical stimulation stress.

Purification and characterization of lead-induced zinc thionein in the liver of rats.

Metallothioneins are involved in homeostasis of essential metals, but its exact biological function remains obscure. Recently it was observed that the administration of lead increased zinc content in the low molecular-weight protein fraction in the rat hepatic 105000x g soluble fraction by using Sephadex G-75 gel filtration, and that this fraction contained only zinc, not lead, as metal. Therefore, the authors have investigated the effects of some drugs on this zinc contained low molecular-weight protein (zinc binding protein, Zn-BP), and further isolated and purified the ZN-BP from liver of rats treated with lead.

Partitioning of copper among copper-binding proteins in the mussel Mytilus edulis exposed to soluble copper

Partitioning of copper among copper-binding proteins was evaluated in digestive glands of Mytilus edulis exposed to soluble copper. Groups of mussels were held in flow-through bioassay systems and exposed to either approx. 1 (control) or 25 ..mu..g Cu/L for as long as 21 weeks. At 3-week intervals, groups of 25 mussels were removed and the digestive glands were analyzed for copper-binding proteins by gel-permeation chromatography and atomic absorption spectrometry. Chronic exposure to copper resulted in increased amounts of copper in the low molecular-weight (LMW) protein fraction, which contains metallothionein-like proteins, and in the high molecular-weight (HMW) protein fraction, which contains metalloenzymes. Concentrations of copper in the LMW protein fraction increased and then appeared to plateau with long exposure times, whereas those in the HMW protein fraction continued to increase with exposure time.

Proteinuria in intact and splenectomized dogs after running and swimming.

The occurrence of post-exercise proteinuria was investigated in intact and splenectomized dogs after treadmill running and swimming and compared to control experiments. Albumin and lysozyme were measured by radial diffusion. Urinary protein was analyzed by SDS-polyacrylamide gel electrophoresis. Swimming in the splenectomized dogs increased the albumin excretion in the first 30 min after exercise from 0.03 to 0.22 mg X min-1 and the lysozyme excretion in the same period from 0.11 to 0.75 micrograms X min-1. Swimming in intact dogs caused smaller increase in the lysozyme and albumin excretions during the exercise period itself as well as in the albumin excretion in the first 30 min after exercise. Running had no effect on urinary albumin or lysozyme but increased the low molecular weight protein fraction in the splenectomized dogs. Plasma lactate concentrations were higher during swimming in the splenectomized dogs than in the intact dogs. Possible mechanisms of post-exercise proteinuria are discussed.

Ligands binding cadmium, zinc, and copper in a species of New Zealand oyster (Ostrea lutaria).

Analysis of Cd, Zn and Cu in the New Zealand oyster, Ostrea lutaria, showed that on an average it contained 5.8 ..mu..g Cd, 67 ..mu..g Zn and 14.1 ..mu..g Cu per g wet weight. Within the oyster tissue, the amount of Cd was equally distributed between the particulate and soluable (cytosol) fractions. The distribution of Zn and Cu was different, about 70% and 80% respectively being present in the cytosol. Separation of the Cd-binding ligands in the oyster cytosol on Sephadex G-75 column showed that about 60% of the cytosolic Cd was bound to high molecular weight (HMW) proteins eluted near the void volume. Similarly about 40% of Zn and Cu was also present in the HMW proteins.A significant proportion (32%) of the cytosolic Cd was present in low molecular weight (LMW) protein fractions. The protein was estimated to have a molecular weight in the range 6000 to 12,000 daltons and the protein fraction was defined as the metallothionein fraction. Only a very small amount (<1%) of Zn was associated with the MT fraction to the high concentration of Cu (2.23 ..mu..g Cu/g wet tissue). The LMW fraction of the oyster tissue cytosol had less than 1% of cytosolic Cd,more » but it contained a large proportion of Zn (about 60%) and Cu (about 32%).« less

Time course of gold-induced accumulation of copper and zinc and the effects of dimercaptosuccinate and cadmium on gold metabolism in rat kidney

After the s.c. administration of sodium aurothiomalate (SATM, 10 mg Au/kg) to male rats, the gold content of the kidney increased to a maximum after 4 days and thereafter declined slowly. The total copper content of the kidney increased at least until 11 days after SATM treatment but was not simply a function of the renal gold content. Gold and copper accumulated in the metallothionein-like, low molecular weight protein fraction and the initial uptake of gold by this fraction appeared to be related to the accumulation of copper. The Zn 2+ content of the kidney was initially unchanged but later increased with the additional Zn 2+ bound predominantly to the non-soluble components. The accumulation of low molecular weight protein-bound copper was not related either to the Zn 2+ content of this fraction or to the total Zn 2+ content of the kidney. Daily administration of dimercaptosuccinic acid (DMSA, 50 mg/kg, i.p.) for 2 weeks to SATM-pretreated rats resulted in a 50% reduction in the concentration of gold in the non-soluble fraction and in both the high and low molecular weight protein components of the soluble fraction. Copper, which accumulated in the non-soluble components and in the soluble low molecular weight protein, was lost only from the non-soluble components. Uptake of gold by the kidneys was increased when rats were pretreated with Cd 2+. The accumulation of gold in the low molecular weight protein fraction was also increased, but most of the additional gold in the kidneys was accumulated by the non-soluble components. SATM had no effect on the renal concentration or subcellular distribution of Cd 2+. It was concluded that the soluble low molecular weight metal binding protein does not have prominent regulatory or protective functions in the renal metabolism of gold.

Hepatic Metallothionein in Rainbow Trout (Salmo gairdneri) as an Indicator of Metal Pollution in the Campbell River System

Hepatic metallothionein was measured in livers of freshly killed rainbow trout (Salmo gairdneri) using differential pulse polarography. The fish were caught in metal-contaminated lakes of the Campbell River watershed and in a nearby control lake. The livers were analyzed for zinc, copper, and cadmium using atomic absorption spectrophotometry. High and low molecular weight protein fractions were separated by gel chromatography from liver cytosols and analyzed for metals. A downward trend from the most contaminated lake to the least was found in levels of zinc in the water, of cadmium and copper in high molecular weight fractions, and of copper in low molecular weight fractions and metallothionein. The concentration of metallothionein is a useful quantitative measure of the degree of exposure of fish to heavy metals.Key words: metallothionein, rainbow trout, Salmo gairdneri; heavy metal pollution, sublethal exposures, mine wastes

Induction of Zinc Metallothionein in the Liver of Rats by Lead

At 18 h after the single injection of lead acetate (100 mg/kg) in male Wistar rats, zinc content in the low molecular-weight protein fraction (zinc binding protein, Zn-BP) in the hepatic 105000g soluble fraction markedly increased, whereas lead was detected only in the high molecular-weight protein fraction by using Sephadex G-75 gel filtration. The properties of this Zn-BP were investigated and the following results were obtained. 1) Molecular weight was estimated about 10000 daltons. 2) Zinc in Zn-BP was substituted with cadmium. 3) UV spectrum of Zn-BP showed similar spectrum to metallothionein. 4) The formation of Zn-BP induced by lead was almost inhibited by the pretreatment with actinomycin D. 5) Zn-BP was heat-stable, when treated at 80°C for 1 min. These findings suggest that Zn-BP is thought to be zinc metallothionein (Zn-MT) and consequently lead has an inductive potency to Zn-MT in the liver of rats.

Assimilatory sulfur metabolism in marine microorganisms: Sulfur metabolism, protein synthesis, and growth of Pseudomonas halodurans and Alteromonas luteo-violaceus during unperturbed batch growth

Analysis of the distribution of 35S-sulfate and 14C-glutamate in major biochemical components of the two marine bacteria, Pseudomonas halodurans and Alteromonas luteo-violaceus, was compared with cell density and total cellular protein during exponential growth in batch culture. For both organisms, the sulfur distribution was restricted principally to the low molecular weight organic and protein fractions, which together accounted for over 90% of the total sulfur. Carbon was more widely distributed, with these two fractions containing only 70% of the total label.

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.

Studies on immunoreactive ACTH from human term placenta. (I) Detection of a high molecular weight-immunoreactive ACTH in term placenta (author's transl)

Extracts of human term placenta were fractionated by Sephadex G-75 gel filtration and assayed for immunoreactive ACTH. Both high and low molecular weight protein fractions were detected to be immunologically reactive toward anti-human ACTH (1--39 alpha) antibody. For the extraction of low molecular weight ACTH from human term placenta (pl. -ACTH), a glacial acetic acid-acetone mixture was employed, while a pH 3.0-HCl solution was used for high molecular weight immunoreactive ACTH. The high molecular weight immunoreactive ACTH fraction (F-I), co-eluted with horse hemoglobin from a Sephadex G-75.column in 0.1M acetic acid, was essentially devoid of low molecular weight materials as revealed by polyacrylamide gel disc electrophoresis at pHs 9.5 and 4.3. Tryptic digestion of F-1 at pH 8.1 and 37 degrees C for 4 hr with E/S of 1/100, followed by fractionation with a Sephadex G-75, resulted in the formation of lower molecular weight fragments. One fragment was eluted at the same position as that of porcine ACTH with a recovery of 86% of immunoreactivity of F-I. Another fragment which was eluted last exhibited positive beta-endorphin receptor binding activity. These results suggest the presence of a common precursor protein to ACTH and beta-endorphin in human term placenta.

Lipoprotein-elastin interactions in human aorta fibrous plaque lesions

The nature of lipoprotein and elastin associated with hyaluronate in human atherosclerotic plaque tissue was studied by digesting the tissues with elastase. The elastase-solubilized lipoprotein—hyaluronate complexes, isolated by Bio-Gel A-50m column chromatography, contained 7.8 mg calcium/100 mg protein. The Sephadex G-200 chromatography of delipidated complexes yielded high (fraction I) and low (fraction II) molecular weight protein fractions. Dialysis of the complexes against 0.01% EDTA resulted in removal of fraction II. Fraction I reacted immunologically against antihuman LDL, and its amino acid composition resembled that of human apoB. Fraction II contained 5 peptide fragments and had high levels of nonpolar amino acids that are characteristic of elastin. However, these peptides also contained high levels of polar amino acids, thus resembling the plaque elastin. These findings suggest that certain regions of plaque elastin may have affinity for apoB containing lipoproteins and calcium.

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.

Atomic absorption spectrometric determination of the total content and distribution of chromium in blood serum

Various atomic absorption spectrometric methods, including the solid sampling technique, were used to determine the total content of chromium in a lyophilized reference sample of animal serum. The concentration in the solid sample was found to be 61 ppb. The distribution of the element among the serum proteins was studied by gel filtration and atomic absorption spectroscopy. Chromium was found to be concentrated in the high and low molecular weight protein fractions.

Entamoeba histolytica: Isolation of ferredoxin

A significant portion of the nonheme iron and acid-labile sulfide of axenically grown Entamoeba histolytica was obtained in a colored, low molecular weight protein fraction from centrifuged amoeba homogenates following gel filtration. The colored component was further purified by adsorption on and elution from DEAE-cellulose. Preparations so purified had 390 280 absorbance ratios as great as 0.87. The oxidized form had absorbance peaks at 390, 315, and 280 nm; the enzymatically reduced form, at 350 nm. The difference spectra showed a major peak at 420 nm, and a minor one at 310 nm. The substance was converted to apoprotein and reconstituted with iron and H 2S by methods developed for clostridial ferredoxin. When chromatographed on a Sephadex G-50 column calibrated with spinach and clostridial ferredoxins its molecular weight appeared to be about 6000, i.e., close to that of the clostridial protein. The substance is considered to be amoebal ferredoxin, but there remains some uncertainty concerning the number of atoms of iron and acid-labile sulfide per molecule. Its physiological role in the organism is not yet understood.