Amino Acid Composition Of Fractions Research Articles

Production and characterization of an extensive rapeseed protein hydrolysate

AbstractRapeseed protein isolate has been used as starting material for the generation of an extensive protein hydrolysate. Protein hydrolysis was produced by using sequentially an endopeptidase (Alcalase) and an exopeptidase (Flavourzyme). The final hydrolysate has a 60% degree of hydrolysis and was completely soluble between pH values 2.5 and 7. Molecular weight profile of the protein hydrolysate was characterized by gel filtration chromatography. A reduction in protein size was observed during the hydrolysis process with accumulation of small peptides and free amino acids after Flavourzyme digestion. Amino acid composition of fractions with different molecular weights of the final hydrolysate was analyzed. Some of these fractions, enriched or poor in certain amino acids, could be used for supplementation or treatment of determined clinical syndromes.

AMINO ACID ANALYSIS OF BEER POLYPEPTIDES

The principles of amino acid analysis of proteins and polypeptides are reviewed. Analysis of the amino acid composition of dialysed beer material prepared from a wide variety of commercial and pilot brewery beers showed that the principal amino acids comprised glutamic acid/glutamine, proline, glycine and aspartic acid/asparagine. The results from the analysis of a series of pilot brewery beers brewed under standardised conditions showed that the composition of the grist may influence the amino acid composition of beer polypeptide fractions. Dialysed beer material prepared from beer brewed from grists containing torrified wheat, wheat flour and malted wheat contained greater proportions of glutamic acid/glutamine compared to material prepared from all malt beers. Further fractionation and analysis of dialysed beer material prepared from pilot brewery beers suggested that fractions MW>60000 contained polypeptide material derived from yeast mannan-protein. In addition fractions MW>60000 prepared from beer brewed from grists containing torrified wheat, wheat flour or all malted wheat may contain high molecular weight polypeptide material derived from wheat proteins. The results from the analysis of fraction MW 40,000–60000 prepared from beers brewed from grists containing all malt, 80% malt and 20% torrified wheat and 50% malt and 50% malted wheat are consistent with the presence of polypeptide material derived from cereal albumins and globulins whereas fractions MW 40,000–60000 prepared from beers brewed from 80% malt and 20% wheat flour and 100% malted wheat may contain polypeptide material derived from wheat prolamins and glutelins. The amino acid composition of fraction MW 20,000–40,000 from all pilot brewery beers investigated is consistent with the presence of polypeptide material derived from cereal prolamins and glutelins. The amino acid composition of beer polypeptide fractions may be used to detect the use of wheat adjuncts in beer brewing.

High-performance liquid chromatography of reduced glutenin: amino acid composition of fractions and components.

Reduced glutenin is separated by gel permeation high-performance liquid chromatography into three major and five minor fractions, which significantly differ in their amino acid compositions. By reversed-phase high performance liquid chromatography, about 20 glutenin components are obtained. These can be classified into three groups according to their amino acid compositions: a hydrophilic group with relatively high values of Glx and Phe, a more hydrophobic group with a high content of Gly, and a strongly hydrophobic group with higher values of Val and Leu. Groups 1, 2 and 3 contain middle-, high- and low-molecular-weight (MMW-, HMW-, LMW-) subunits respectively.

Amino acid composition of fractions of ‘Kentucky-31’ tall fescue as affected by N fertilization and mild water stress

Environmental and management factors can influence the protein concentration of forages, significantly altering specific amino acid content. Drought, high rates of fertilizer N and the presence of a fungal endophyte have been associated with significant alterations in plant N metabolites and animal performance problems on tall fescue. A controlled environment study was conducted to examine the influence of N fertilization (10 and 100 μgN/g) and water regime (low and adequate soil water availability) upon the distribution and concentration of amino acids in endophyte infected tall fescue (Festuca arundinacea, Schreb.) herbage. Tall fescue tissue was collected from three replicates of each treatment, quick frozen in liquid N and lyophilized. Two insoluble (RI, structural residue; RII, membrane residue) and two soluble (SI, soluble protein; SII, low molecular weight N compounds) fractions were collected. Amino acid analyses of acid hydrolysates of fractions showed that application of 100 N significantly increased the concentration (per unit dry weight) of all amino acids in the entire plant, with an average increase of about 55%. Application of 110 N increased the concentrations of most amino acids in fractions RI, RII, and SI, but only aspartate-asparagine, glutamate-glutamine, alanine, threonine, serine, valine and proline in fraction SII. Fraction RI contained about 65% of total amino acids under 10 N and 55% under 110 N even though N level did not alter dry matter distribution among fractions. While the amount of dry matter was least in SI, amino acids in the fraction ranged from 8% (leucine, 10 N) to 20% (lysine, 110 N) of the total amount of specific amino acids recovered. Significant increases in proline, glutamate, aspartate, serine, valine, threonine, alanine and phenylalanine concentration occurred under low soil-water availability compared with adequate water conditions. Basic amino acids including histidine, arginine and lysine increased with increased N and with water stress at each N level. Application of N increased amounts, and water stress influenced distribution of amino acids among the fractions of tall fescue herhage. Nitrogenous components, such as non-protein amino acids which could influence plant nutritive quality, were increased in fraction SII by increased N and water stress.

Storage Proteins of Glandless Cottonseed Flour

ABSTRACTThe salt‐soluble storage protein isolate of defatted glandless cottonseed flour was separated by gel filtration column chromatography into six fractions (I to VI). Column and thin‐layer gel filtration showed that these fractions had molecular weights of >600,000, 280,000, 127,000, 63,500, 10,700, and <2,000, respectively. Treating fraction I protein with dilute alkali dissociated a component that migrated upon gel filtration with fraction VI. Both fractions I and VI were yellow and the color intensified at alkaline pH, suggesting the presence of bound flavonol and gossypol pigments. Because of its large size and low (16.6%) protein content, fraction I was thought to be fragments of membrane and aleurone gram globulins. Gradient polyacrylamide gel electrophoresis (PAGE) showed that the proteins in each fraction being completely recovered from column chromatography were represented in the pattern of the initial isolate. The proteins in fractions II and III have similar ammo acid compositions, except that III is low in methionine, cystine and tryptophan and II is low in tyrosine and phenylalanine. The amino acid composition of fraction I was also similar to fractions II and III, while unique patterns were noted for fractions V and VI. Fraction VI was rich in lysine and arginine.

EFFECT OF NITROGEN FERTILIZER ON NITROGEN FRACTIONS AND AMINO ACID COMPOSITION OF SPRING WHEAT

Grain from hard red spring wheat (Triticum aestivum ’Neepawa’) grown with nine rates of N fertilizer (0–400 kg/ha) was analyzed for total N, non-protein N (NPN), and amino acids. In addition, the amino acid compositions of the various soluble protein fractions were determined. The percentages of total N, NPN, and protein N of the grain increased with each 50-kg increment of N up to the 150-kg rate. The proportions of glutamate, proline, and phenylalanine in the seed increased whereas threonine, serine, glycine, alanine, and valine decreased with the first three or four increments of N fertilizer. With increasing N content in the seed, the proportions of albumin, globulin, and glutenin 2 decreased while the proportions of gliadin and glutenin 1 increased. The various protein fractions differed from each other in amino acid composition but the amino acid composition of each fraction was not influenced significantly by N application.

Heparin-sepharose affinity chromatography of human post-heparin plasma. Characterization of the elution pattern with immunoelectrophoretic methods.

Immunochemical methods including fused rocket-, crossed-, and tandem-crossed immunoelectrophoresis, have been used to characterize the elution pattern from heparin-Sepharose affinity, chromatography of human post-heparin plasma. Hepatic triglyceride lipase (H-TGL) but not lipoprotein lipase (LPL) could be visualized with beta-naphthyl acetate after immunoelectrophoresis. Two proteins were found to elute together with the lipolytic enzymes. The amino acid composition of fractions containing these proteins was nearly identical to that of antithrombin III. These results indicate that the removal of antithrombin III is the major problem in the purification of H-TGL and LPL from human post-heparin plasma by heparin-Sepharose affinity chromatography.

Isolation and partial characterization of the major apolipoprotein component of bovine serum high density lipoprotein

The delipidated protein component of bovine serum high density lipoprotein was fractionated by gel filtration on a Sephadex G-150 column (equilibrated with buffer containing 6 M urea) into three fractions: I, II and III. Fractions I and II together constitute 88% of all the protein weight of bovine high density lipoprotein, whereas fraction III accounts for the remaining 12%. Analysis of the fractions by sodium dodecyl sulfate-polyacrylamide electrophoresis reveals that fraction I consists mostly of aggregated forms of fraction II and some higher molecular weight species, probably irreversible aggregates of fraction II. The irreversible aggregates are apparently formed during the delipidation procedure or upon aging of the lipoprotein. The major protein component of the high density bovine lipoprotein is found in fraction II; it has a molecular weight of 27 000 ± 1500 and appears to be homogeneous by several physicochemical criteria. The amino acid composition of fractions I and II are essentially identical; their spectral properties, including absorption, fluorescence, and circular dichroism spectra, are similar; however, fraction I appears to contain traces of oxidized lipid and more secondary, α-helical organization than fraction II. By comparison with the intact lipoprotein, which contains about 65% of α-helical structure, fractions I and II have diminished α-helical organization, 55% and 43%, respectively. Fraction III, on sodium dodecyl sulfate-polyacrylamide electrophoresis, separates into two protein bands of equivalent intensity, having molecular weights around 13 000 and 11 000. Fraction III is markedly distinct from the other two, in amino acid composition and spectral properties, especially in its red-shifted fluorescence and very low content of α-helical structure. The protein composition of bovine serum high density lipoprotein is compared with recently published results for high density lipoprotein apoproteins of man, chimpanzee, rhesus monkey, pig and rat. Similarities and differences are discussed in terms of possible evolutionary and functional factors.

The amino-acid compositions of CNBr fragment C1 from antihapten antibodies. Use of guanidine for reproducible isolation of the C1 fragment.

Most rabbit antibodies contain a methionine residue about 240 residues from the NH2‐terminus of the heavy chain sequence. Cleavage of heavy chains by CNBr at this methionine leads to the formation of a fragment, C1, which contains the variable portion of the H‐chain but is slightly longer than the Fd fragment. Published amino acid compositions of fractions from different antibodies, all taken to represent fragment C1, showed marked similarity. In this work it is shown that the primary reason for this similarity is that previous methods of isolating C1, by gel filtration using 6 M urea or 1 M acetic acid as solvent, led to the isolation of aggregates contaminated with an extensive section of the constant region. Six M guanidine · HCl prevented the formation of these aggregates and gel filtration in this solvent yielded a C1 fragment with an amino acid composition quite different from that of the contaminated aggregate. The amino acid compositions of the uncontaminated C1 fragments from antihapten antibodies from individual rabbits showed large differences. The differences were apparent because of the restricted heterogeneity of the antibodies being compared. Amino acid compositions of C1 fragments from γ‐globulins from individual uninjected rabbits showed appreciably less variation because, unlike the antibodies, the γ‐globulins were complex mixtures.

Distribution of carbohydrate among the polypeptide chains and plasmin digest products of human fibrinogen

Human fibrinogen was subjected to sulfitolysis and the constituent polypeptide chains were isolated. Each of the three chain types contained glucosamine and hexose (analyses for sialic acid were not performed). The glucosamine and hexose contents of fibrinogen were satisfied by the sum of the corresponding values for a pair of each of the three chains. Pronase digest glycopeptides from the three chains were compared with those from three fractions of a prolonged plasmin digest of fibrinogen (designated fractions D, E, and F). Fraction D of about 50,000 mol wt yielded a glycopeptide fraction which was similar to that from the α(A)-chain. Fraction E of about 30,000 mol wt contained the glycopeptide portion of the γ-chain, while the glycopeptide portion of the β (B)-chain was contained in a heterogeneous low molecular weight fraction, here designated F. The values for molecular weight, glycopeptide composition, and amino acid composition of fraction E resembled those for the “disulfide knot” of fibrinogen which contains the N-terminal portions of the three chains.

Radiation chemistry of proteins. I. Modified lysozyme substances

Crystalline egg-white lysozyme has been irradiated with 0.66 m.e.v. γ-rays, and the resulting sample has been subjected to amino acid analyses, fractionation by phosphate precipitation, and column chromatography on Bio-Rex 70. No significant changes are observed in amino acid levels in samples given a 30 Mrads dose in the presence of air. In samples receiving 29.8 Mrads in vacuo, no significant changes occur in amino acid composition of fractions obtained by phosphate precipitation. In samples receiving 124.8 Mrads in vacuo, the phosphate-soluble fraction contains appreciably larger numbers of lysine, arginine, and valine residues per 131 peptide link ages than the unirradiated lysozyme. Chromatographic elution patterns of phosphate-soluble fractions of irradiated lysozyme samples reveal several enzymically active proteins. Four of the Chromatographic peaks retain enzymic activity. At levels of 14.5 and 71.5 Mrads the main peak is higher in enzymic specific activity than the original material and is eluted earlier from the column. Deuterium-exchange rate studies show that peptide hydrogens in the protein are more susceptible to exchange with deuterium after irradiation. Hard-to-exchange amide (peptide) hydrogens are lost at least twice as rapidly as enzymic activity as a function of radiation dose up to 45 Mrads.

Amino Acid Composition of Normal Wools, Wool Fractions, Mohair, Feather, and Feather Fractions

Amino acid analyses of normal wools of several breeds and geographical origins are presented as evidence of the normal variation of composition. Some samples were from lots used for detailed comparative studies of fiber properties and processing behavior. A mohair sample was found to be similar to wool in composition. White Leghorn chicken feather keratin differed from wool in yielding larger amounts of serine and proline. Feather had characteristically smaller amounts of cystine and glutamic acid. Heterogeneity of wool and feather keratins was shown by differences in amino acid composition of fractions prepared by treatment with acid, by mechanical separation, or by partial solubilization in aqueous alcohol. Feather differs from wool in having readily separable fractions differing greatly in content of histidine, lysine, and methionine. These fractions also differ appreciably in their content of several other amino acids.