scholarly journals The polypeptide composition of bovine epidermal α-keratin

1975 ◽  
Vol 151 (3) ◽  
pp. 603-614 ◽  
Author(s):  
P M Steinert ◽  
W W Idler
Keyword(s):  
Amino Acid ◽  
Glutamic Acid ◽  
Polyacrylamide Gel Electrophoresis ◽  
Deae Cellulose ◽  
Cysteine Residues ◽  
Net Charge ◽  
Extinction Coefficients ◽  
Molecular Weights ◽  
Α Helix ◽  
Polypeptide Chains

1. The polypedtide chains that comprise the subunits of the tonofilaments, or the α-keratin component, of bovine epidermis were fractionated by combination of chromatography on DEAE-cellulose and preparative polyacrylamide-gel electrophoresis. 2. The seve polypeptide chains investigated had generalyy similar properties; all contained two residues per molecule of tryptophan and N-acetylserine was the common N-terminal amino acid residue. 3. On the basis of close similarities in α-helix content and amino acid composition, the polypeptide chains were classified into three distinct groups. Each group contained approximately one-third of the total polypeptides on a molar basis. The groups and designated polypeptides chain numbers were: group one, polypeptides 1a and 1b, which had moleculae weights of 58,000, contained about 25% α-helix, 86 glutamic acid and 8 cysteine residues per molecule, but which differed in net charge, extinction coefficients and tyrosine contents; group two, polypeptides 2, 3, and 4, which hadmolecular weights within thewithin the range of 52,00-56,000, contained about 48% α-helix, 54 glutamic acid and 6 cysteine residues per molecule, but which differed in extinction coefficients and tryosine contents; and group, polypeptides 5 and 6, which had molecular weights of 47000-48000, contained about 56% α-helix, 64 glutamic acid and 4 cysteine residues per molecule, but which differed in extinction coefficients and tyrosine contents, it is suggested that none of the chains is a precursor or a degradation product of other polypeptidc chains. 5. It is concluded that bovine epidermal α-keratin consists of a heterogeneous group of similar polypeptide chains.

scholarly journals Isolation and characterization of phytoferritin from pea (Pisum sativum) and Lentil (Lens esculenta)

1978 ◽  
Vol 171 (2) ◽  
pp. 349-356 ◽  
Author(s):  
R R Crichton ◽  
Y Ponce-Ortiz ◽  
M H J Koch ◽  
R Parfait ◽  
H B Stuhrmann
Keyword(s):  
Amino Acid ◽  
Pisum Sativum ◽  
Quaternary Structure ◽  
Polyacrylamide Gel Electrophoresis ◽  
Lens Esculenta ◽  
Molecular Weights ◽  
Isolation And Characterization ◽  
Angle Scattering ◽  
Scattering Study ◽  
Polypeptide Chains

Ferritin was isolated from the seeds of pea (Pisum sativum) and lentil (Lens esculenta). The homogeneity of the phytoferritins was established by polyacrylamide-gel electrophoresis. The subunit molecular weights were respectively 20 300 and 21 400 for hte pea and lentil proteins. A neutron low-angle scattering study established the molecular weight of the oligomer as 480 000 for pea apoferritin and 510 000 for lentil apoferritin. Although the quaternary structure of 24 polypeptide chains is preserved, the phytoferritins have a larger cavity in the interior than mammalian ferritins and can thus potentially store 1.2-1.4 times as much iron. The amino acid composition of the phytoferritins show some similarities to those of mammalian apoferritins; tryptic ‘fingerprinting’ reveals that there are many differences in the amino acid sequence of plant and mammalian apoferritins.

scholarly journals Chemical properties of guinea-pig immunoglobulins γ2G and γM

1970 ◽  
Vol 120 (4) ◽  
pp. 787-795 ◽  
Author(s):  
R. G. Q. Leslie ◽  
S. Cohen
Keyword(s):  
Amino Acid ◽  
Guinea Pig ◽  
Chemical Properties ◽  
Biological Properties ◽  
Extinction Coefficients ◽  
Amino Acid Compositions ◽  
Molecular Weights ◽  
Structural Differences ◽  
Polypeptide Chains

The isolation of guinea-pig immunoglobulins γ1G, γ2G and γM are described and methods for separating the polypeptide chains of each examined. The molecular weights, extinction coefficients and carbohydrate and amino acid compositions of the immunoglobulins and their constituent chains have been analysed. The findings provide a basis for further studies attempting to relate structural differences to distinct biological properties of guinea-pig immunoglobulins.

scholarly journals α-Crystallin. The isolation and characterization of distinct macromolecular fractions

1971 ◽  
Vol 124 (2) ◽  
pp. 337-343 ◽  
Author(s):  
Abraham Spector ◽  
Lu-Ku Li ◽  
Robert C. Augusteyn ◽  
Arthur Schneider ◽  
Thomas Freund
Keyword(s):  
Molecular Weight ◽  
Amino Acid ◽  
Gel Filtration ◽  
Deae Cellulose ◽  
Chemical Difference ◽  
Molecular Weights ◽  
Isolation And Characterization ◽  
Different Populations ◽  
Molecular Weight Fractions

α-Crystallin was isolated from calf lens periphery by chromatography on DEAE-cellulose and gel filtration. Three distinct populations of macromolecules have been isolated with molecular weights in the ranges approx. 6×105−9×105, 0.9×106−4×106and greater than 10×106. The concentration of macromolecules at the molecular-weight limits of a population are very low. The members of the different populations do not appear to be in equilibrium with each other. Further, in those molecular-weight fractions investigated, no equilibrium between members of the same population was observed. The population of lowest molecular weight comprises 65–75% of the total material. The amino acid and subunit composition of the different-sized fractions appear very similar, if not identical. The only chemical difference observed between the fractions is the presence of significant amounts of sugar in the higher-molecular-weight fractions. Subunit molecular weights of approx. 19.5×103and 22.5×103were observed for all α-crystallin fractions.

Subunit studies of coupling factor 1 of bean chloroplasts

1976 ◽  
Vol 54 (5) ◽  
pp. 481-487 ◽  
Author(s):  
M. P. Silvanovich ◽  
R. D. Hill
Keyword(s):  
Amino Acid ◽  
Gel Electrophoresis ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Coupling Factor ◽  
Leaf Extracts ◽  
Molecular Weights ◽  
Major Species ◽  
Chloroplast Coupling Factor ◽  
Coupling Factors

A bean chloroplast coupling factor (CF1) with latent Ca2+-dependent ATPase activity was studied. Immunodiffusion of bean (Phaseolus vulgaris) chloroplast and etioplast coupling factors and spinach coupling factor against antiserum to spinach coupling factor showed partial identity of the bean coupling factor with that of spinach. An immunoelectrophoretic comparison, under dissociating conditions, of bean leaf extracts and spinach extracts containing CF1 subunits (as determined by sodium dodecyl sulfate polyacrylamide gel electrophoresis) gave identical results for both extracts. At least six distinct polypeptide species were found. The major species had molecular weights of 42 000, 59 000 and 63 000 daltons. Amino acid analysis of electrophoretically purified bean CF1 gave results similar to those published for spinach CF1.

Existence of Common Antigenic Determinants in the Aα, Bβ and γ Chains of Some Mammalian Fibrinogens.

1979 ◽  
Author(s):  
C.S. Cierniewski
Keyword(s):  
Amino Acid ◽  
Polyacrylamide Gel Electrophoresis ◽  
Polyacrylamide Gel ◽  
Antigenic Determinants ◽  
Terminal Amino Acid ◽  
Human Fibrinogen ◽  
Passive Hemagglutination ◽  
Terminal Amino ◽  
Polypeptide Chains ◽  
Fibrinogen Molecule

Polypeptide chains Aα, Bβ and γ of porcine fibrinogen were isolated by preparative SDS polyacrylamide gel electrophoresis. Their purity was estimated by electrophoresis in polyacrylamide gel, amino acid composition and N-terminal amino acid analyses. Antisera to the pig polypeptide chains were produced in rabbits and they were employed in immunological comparative studies of porcine, bovine, human and duck fibrinogens. Antisera to the pig Aα chain showed in gel immunodiffusion and passive hemagglutination a strong cross-reaction with porcine, bovine and human fibrinogens. Antisera to the pig βB and γ chains cross-reacted only with porcine and bovine fibrinogens but they did not recognize human fibrinogen, The reaction of antiγ antisera was detectable only by passive hemagglutination test. In all cases antigenic similarity of the analyzed fibrinogens was mainly related to antigenic determinants of the Aα, Bβ and γ chains exposed on the intact fibrinogen molecule. None of analyzed antisera reacted with duck fibrinogen.

scholarly journals α-Crystallin. Fractionation of subunits and sequence studies on an isolated polypeptide

1971 ◽  
Vol 124 (2) ◽  
pp. 345-355 ◽  
Author(s):  
Robert C. Augusteyn ◽  
Abraham Spector
Keyword(s):  
Amino Acid ◽  
Gel Electrophoresis ◽  
Cyanogen Bromide ◽  
Sodium Dodecyl ◽  
Deae Cellulose ◽  
Iodoacetic Acid ◽  
Cysteine Residue ◽  
Terminal Sequence ◽  
Amino Acid Compositions ◽  
Molecular Weights

α-Crystallin was carboxymethylated with radioactive iodoacetic acid in the presence of 7.6m-urea and then separated into six major fractions by chromatography on DEAE-cellulose in 7m-urea. Based on the amino acid compositions, specific radioactivities and sodium dodecyl sulphate–gel electrophoresis of the fractions, it was concluded that α-crystallin contains at least four different subunits: DU1A and DU1B, containing no cysteine; a third component represented by DU2B and DU3 containing one cysteine one cysteine residue per subunit; and DU4, which probably contains two residues of cysteine per subunit. Subunit DU1A was shown to be of sufficient purity for sequence studies. Cyanogen bromide cleavage yielded two peptides, CB-1 and CB-2, in approximately equal amounts as expected. The sum of the molecular weights and amino acid compositions of the peptides were both in excellent agreement with the results obtained for subunit DU1A. The amino acid sequence of the first sixteen residues of peptide CB-1 is: Ser-Leu-Thr-Lys-Asp-Phe-Asp-Glu-Val-Asn-Ile-Asp-Val-Ser-His-Phe-. The sequence of the first seventeen residues of peptide CB-2 is: Asp-Ile-Ala-Ile-Ser-His-Pro-Trp-Ile-Arg-Pro-Ser-Phe-Phe-Glu-Phe-His-. The N-terminal sequence of subunit DU1A was shown to be N-acetylmethionine followed by peptide CB-2.

scholarly journals Biochemical characterization of a second family of human Ia molecules, HLA-DS, equivalent to murine I-A subregion molecules.

1982 ◽  
Vol 156 (2) ◽  
pp. 550-566 ◽  
Author(s):  
S M Goyert ◽  
J E Shively ◽  
J Silver
Keyword(s):  
Amino Acid ◽  
Amino Acid Sequence ◽  
Amino Acid Sequence Identity ◽  
Biochemical Characterization ◽  
Amino Acid Sequence Homology ◽  
Molecular Weights ◽  
Hla Dr ◽  
D Region ◽  
Polypeptide Chains

In mice, two families of structurally distinct Ia molecules, one designated I-A and the other I-E, have been identified and characterized. The HLA-DR molecules represent one family of human Ia molecules equivalent to the murine I-E molecules on the basis of amino acid sequence homology. We describe the isolation and biochemical characterization of a second family of human Ia molecules, designated HLA-DS for second D-region locus, equivalent to the murine I-A molecules. The human HLA-DS molecules consist of two polypeptide chains, DS alpha (37,000 mol wt) and DS beta (29,000 mol wt), with 73% amino acid sequence identity to the murine I-A molecules. Furthermore, the HLA-DS molecules are closely linked genetically to HLA-DR molecules, a situation analogous to that observed in mice. The similarity in molecular weights of the DR and DS molecules might explain why others have failed to identify the latter in man.

Macrozin, a Novel Storage Globulin From Seeds of Macrozamia communis L. Johnson

1984 ◽  
Vol 11 (2) ◽  
pp. 69 ◽  
Author(s):  
RJ Blagrove ◽  
GG Lilley ◽  
TJV Higgins
Keyword(s):  
Molecular Weight ◽  
Broad Band ◽  
Sodium Dodecyl ◽  
Optical Rotatory Dispersion ◽  
Molecular Weights ◽  
Neutral Ph ◽  
Cellulose Acetate Membranes ◽  
Dodecyl Sulfate ◽  
Α Helix ◽  
Polypeptide Chains

The isolation, characterization and amino acid composition are reported for macrozin, the major storage globulin found in seeds of Macrozamia communis. Electrophoresis of macrozin on cellulose acetate membranes at neutral pH resulted in a single broad band indicating limited charge heterogeneity. Isoelectric focusing under dissociating and reducing conditions showed this globulin to be composed of a family of polypeptide chains with apparent isoelectric points in the range pH 6.0-7.5. Sedimentation equilibrium studies showed that the main component purified by gel filtration in aqueous buffers at neutral pH has a molecular weight of 260 000 and a sedimentation coefficient S020.w = 10.9 S. This component dissociates in 8 M urea to yield subunits of molecular weight 126 000. Each subunit is composed of disulfide-bonded polypeptide chains of approximate molecular weight 44 000. The apparent molecular weights for the macrozin subunit and its constituent polypeptides were 130 000 and 46 000 from dodecyl sulfate-polyacrylamide gradient gel electrophoresis. The dimeric nature of the main oligomer in aqueous solution was confirmed by crosslinking the subunits with dithiobis(succinimidylpropionate); the presence of three polypeptide chains per subunit is inferred from the molecular weights. Optical rotatory dispersion and circular dichroism measurements suggest that macrozin is devoid of α-helix in its native conformation, but contains some 25% α-helix after incubation with sodium dodecyl sulfate.

scholarly journals Cleavage of bacterial flagellin with cyanogen bromide. Chemical and physical properties of the protein fragments

1969 ◽  
Vol 113 (3) ◽  
pp. 489-499 ◽  
Author(s):  
C. R. Parish ◽  
G. L. Ada
Keyword(s):  
Amino Acid ◽  
Gel Electrophoresis ◽  
Amino Acid Analysis ◽  
Cyanogen Bromide ◽  
Degradation Products ◽  
Polyacrylamide Gel Electrophoresis ◽  
Polyacrylamide Gel ◽  
Gel Chromatography ◽  
Molecular Weights ◽  
Lysine Residues

1. Flagellin, isolated from the flagella of Salmonella adelaide, was shown by various criteria to be a pure protein. It had a molecular weight of about 40000 and contained three methionine, six tyrosine, 11 arginine and 25 lysine residues/mol., of which 11 of the lysine residues were present as ∈-N-methyl-lysine. 2. After treatment of flagellin with cyanogen bromide in formic acid, four main fragments (A, B, C and D) were obtained, with as many as six minor components that represented partial degradation products. The major fragments were estimated by amino acid analysis to have molecular weights of about 18000 for fragment A, 12000 for fragment B, 5500 for fragment C and 4500 for fragment D. Fragments A, B and D, but not fragment C, were recovered pure by gel chromatography as monitored by polyacrylamide-gel electrophoresis. 3. A complex between fragments C and D was also isolated (mol.wt. 10000) after limited oxidation of flagellin by chloramine-t before digestion by cyanogen bromide. After oxidation essentially only two fragments were released from flagellin by cyanogen bromide: the ‘C,D’ complex and a presumed ‘AB’ fragment. 4. The sum of the amino acid analyses of fragments A and B and the ‘C,D’ complex gave residue values that agreed well with the amino acid composition of native flagellin. 5. Fragments A and D contained tyrosine, and ten of the 11 ∈-N-methyl-lysine residues of the molecule were in fragment A. Reaction with [125I]iodide at small extents of substitution showed that, in flagellin, the tyrosine residue of fragment D was more readily substituted than those of fragment A. By contrast, in polymerized flagellin, the tyrosine residues of fragment A were more readily substituted. 6. Treatment of flagellin with carboxypeptidases A and B revealed the C-terminal sequence -Leu-Leu-Leu-Arg. Arginine and leucine were released by carboxypeptidase from the ‘C,D’ complex but not from fragment D, indicating that fragment C was C-terminal. 7. On the basis of the results from amino acid analysis, carboxypeptidase digestion, N-terminal analysis, iodination studies and polyacrylamide-gel electrophoresis, the sequence of fragments in flagellin was considered to be B–A–D–C; in the polymer, fragment A was exposed. It is suggested that methylation of the lysine residues occurred in the organism after flagellin had polymerized.

Purification and characterization of the extracellular α-amylase activity of the yeast Schwanniomyces alluvius

1987 ◽  
Vol 65 (10) ◽  
pp. 899-908 ◽  
Author(s):  
F. Moranelli ◽  
M. Yaguchi ◽  
G. B. Calleja ◽  
A. Nasim
Keyword(s):  
Amino Acid ◽  
Amylase Activity ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Gel Filtration ◽  
Deae Cellulose ◽  
Exchange Chromatography ◽  
Proteinase K ◽  
Ph Optimum ◽  
Sds Page

The extracellular α-amylase activity of the yeast Schwanniomyces alluvius has been purified by anion-exchange chromatography on DEAE-cellulose and gel-filtration chromatography on Sephadex G-100. Sodium dodecyl sulfate – polyacrylamide gel electrophoresis (SDS–PAGE) and N-terminal amino acid analysis of the purified sample indicated that the enzyme preparation was homogeneous. The enzyme is a glycoprotein having a molecular mass of 52 kilodaltons (kDa) estimated by SDS–PAGE and 39 kDa by gel filtration on Sephadex G-100. Chromatofocusing shows that it is an acidic protein. It is resistant to trypsin but sensitive to proteinase K. Its activity is inhibited by the divalent cation chelators EDTA and EGTA and it is insensitive to sulfhydryl-blocking agents. Exogenous divalent cations are inhibitory as are high concentrations of monovalent salts. The enzyme has a pH optimum between 3.75 and 5.5 and displays maximum stability in the pH range of 4.0–7.0. Under the conditions tested, the activity is maximal between 45 and 50 °C and is very thermolabile. Analysis of its amino acid composition supports its acidic nature.

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