scholarly journals RECONSTITUTION OF 7S MOLECULES FROM L AND H POLYPEPTIDE CHAINS OF ANTIBODIES AND γ-GLOBULINS

1964 ◽  
Vol 119 (5) ◽  
pp. 789-815 ◽  
Author(s):  
D. E. Olins ◽  
G. M. Edelman
Keyword(s):  
Molecular Weight ◽  
Propionic Acid ◽  
Disulfide Bonds ◽  
Average Molecular Weight ◽  
Antigenic Structure ◽  
Hybrid Molecules ◽  
Different Types ◽  
Γ Globulins ◽  
Chain Fraction ◽  
Polypeptide Chains

Admixture of separated L and H polypeptide chains of 7S γ-globulins under appropriate conditions has been found to result in the reconstitution of 7S molecules. The chains were mixed in 0.5 N propionic acid and when dialyzed into neutral aqueous buffers interacted to form reconstituted material in greater than 30 per cent yield. This material had sedimentation coefficients of 6S to 7S, a weight average molecular weight of 160,000, and its antigenic structure and electrophoretic properties were the same as those of 7S γ-globulin. By the use of I131 and I125 labels on the different types of chains, combined with ultracentrifugation of chain mixtures in sucrose density gradients, the 7S product was found to contain both isotopes in ratios consistent with the presence of two L and two H chains. After hydrolysis with papain, the reconstituted material yielded products resembling S and F fragments. All of the isotope corresponding to L chains was found in the counterpart of the S fragment; the isotope corresponding to the H chain fraction was present in both fragments. The activity reconstituted from chains of a purified guinea pig antibody to f1 phage was found to be associated mainly with the 7S material. Hybrid molecules containing rabbit L chains and human H chains and of human L chains and rabbit H chains were formed by the same techniques of reconstitution. It was found that the interchain disulfide bonds of native 7S γ-globulins could be broken and reoxidized, as could those of reconstituted 7S material. Reduced L and H chains mixed in propionic acid, dialyzed against neutral buffers containing mercaptan, then against neutral buffers in the absence of mercaptan, formed stable 7S molecules of molecular weight 160,000, which were dissociable only after reduction.

MONOMER AND DIMER FORMS OF GAMMA-GLOBULIN POLYPEPTIDES IN NORMAL URINE

1964 ◽  
Vol 42 (12) ◽  
pp. 1815-1823 ◽  
Author(s):  
M. H. Freedman ◽  
G. E. Connell
Keyword(s):  
Molecular Weight ◽  
Disulfide Bonds ◽  
Gamma Globulin ◽  
The Other ◽  
Light Chains ◽  
Low Molecular Weight ◽  
Γ Globulins ◽  
Intermolecular Disulfide Bonds ◽  
Normal Urine ◽  
Polypeptide Chains

The γ-globulins of low molecular weight in human urine have been partially fractionated and characterized. The two principal components are made up of "light" polypeptide chains of 7 S γ-globulin. One of these components, the "monomer", has a molecular weight in dissociating solvents of approximately 20,000 and probably consists of single light chains. The other component, the "dimer", has a molecular weight of approximately 40,000 and probably consists of pairs of light chains joined by intermolecular disulfide bonds.

scholarly journals STUDIES ON STRUCTURAL UNITS OF THE γ-GLOBULINS

1961 ◽  
Vol 113 (5) ◽  
pp. 861-884 ◽  
Author(s):  
G. M. Edelman ◽  
M. D. Poulik
Keyword(s):  
Amino Acid ◽  
Gel Electrophoresis ◽  
Disulfide Bonds ◽  
Average Molecular Weight ◽  
Structural Features ◽  
Starch Gel Electrophoresis ◽  
Molecular Weights ◽  
Γ Globulins ◽  
Starch Gel ◽  
Polypeptide Chains

When human and rabbit 7S γ-globulins were reduced in strong urea solutions by a number of procedures, their molecular weights fell to approximately ⅓ of the original values. Partial separation of the reduction products was achieved using chromatography and starch gel electrophoresis in urea solutions. One of the components of reduced human 7S γ-globulin was isolated by chromatography, identified by starch gel electrophoresis, and subjected to amino acid analyses. The amino acid composition of this component differed from that of the starting material and also from that of the remaining components. A reduced pathological macroglobulin dissociated to components with an average molecular weight of 41,000. Several reduced human myeloma proteins, when subjected to starch gel electrophoresis, yielded individual patterns that nevertheless had features in common with those of reduced normal γ-globulins. Reduction of normal and abnormal γ-globulins was accompanied by the appearance of titratable sulfhydryl groups. Chemical treatments other than reduction were used to determine the type of bond holding the subunits together. It was tentatively concluded that they were linked by disulfide bonds. An hypothesis is presented to relate the structural features of the various γ-globulins in terms of the multiplicity of polypeptide chains in these molecules.

scholarly journals ACTIVITY OF DISSOCIATED AND REASSOCIATED 19S ANTI-γ-GLOBULINS

1964 ◽  
Vol 120 (6) ◽  
pp. 1215-1229 ◽  
Author(s):  
Ralph E. Schrohenloher ◽  
Henry G. Kunkel ◽  
Thomas B. Tomasi
Keyword(s):  
Rheumatoid Arthritis ◽  
Molecular Weight ◽  
Disulfide Bonds ◽  
Red Cells ◽  
Low Molecular Weight ◽  
Density Gradient Ultracentrifugation ◽  
High State ◽  
Analytical Ultracentrifuge ◽  
Γ Globulins ◽  
Molecular Weight Fractions

19S anti-γ-globulins were isolated in a high state of purity from the sera of two patients with rheumatoid arthritis. Following reduction with ethyl mercaptan and alkylation by iodoacetamide, fragments were produced which retained the capacity to combine with 7S γ-globulin. The fragments from one of the 19S anti-γ-globulins agglutinated red cells coated with incomplete anti-Rh antibodies. This activity was shown by density gradient ultracentrifugation to be associated with low molecular weight fractions. The agglutination of the coated red cells by the fragments was strongly inhibited by normal and myeloma 7S γ-globulins and showed a greater specificity than the parent 19S material. Analytical ultracentrifuge experiments demonstrated that the fragments from either of the 19S anti-γ-globulins formed complexes with 7S γ-globulin. Reassociation of the dissociated fragments through reformation of disulfide bonds resulted in the formation of fast sedimenting molecules having properties similar to those of the untreated 19S material in respect to precipitation with aggregated γ-globulin and agglutination of coated red cells.

scholarly journals CHARACTERISTICS OF AN IMMUNE SYSTEM COMMON TO CERTAIN EXTERNAL SECRETIONS

1965 ◽  
Vol 121 (1) ◽  
pp. 101-124 ◽  
Author(s):  
Thomas B. Tomasi ◽  
Eng M. Tan ◽  
Alan Solomon ◽  
Robert A. Prendergast
Keyword(s):  
Disulfide Bonds ◽  
Defense Mechanisms ◽  
Sedimentation Coefficient ◽  
Antibody Activity ◽  
Local Production ◽  
Local Synthesis ◽  
Distinctive Type ◽  
Γ Globulins ◽  
Polypeptide Chains ◽  
Circulating Antibody

The γ1A present in saliva and colostrum exists largely in the form of higher polymers, the major component of which has a sedimentation coefficient of 11S. The 11S γ1A in these fluids differs from the polymers found in normal and myeloma sera both immunologically and by the fact that their sedimentation coefficients are unaffected by disulfide bond reduction in the absence of urea. However, like other γ-globulins the 11S γ1A molecules consist of multiple polypeptide chains linked by disulfide bonds. Local synthesis of γ1A in the salivary gland has been shown by fluorescent and autoradiographic studies, although the fraction of the total salivary γ1A which is derived from local production is uncertain. No evidence of transport of intravenously administered I131-labeled 7S γ1A from serum to saliva was obtained. Immunological specificity has been demonstrated in the salivary and colostral γ1A. Whether that portion of the γ1A which is immunologically specific is a piece incorporated during the local synthesis of γ1A in the gland or is added by the epithelial cell in the process of transport remains to be determined. Antibody activity (isohemagglutinins) have been demonstrated in saliva and colostrum and have been shown to be of the γ1A-type. In both of these fluids activity is associated primarily with γ1A-polymers of 11S and 18S sizes. There appears to be an immunological system which is characteristic of certain external secretions. Its properties including the local production of a distinctive type of antibody separate it from the "systemic" system responsible for the production of circulating antibody. This system may play a significant role in the body's defense mechanisms against allergens and microorganisms.

scholarly journals THE MITOTIC APPARATUS

1967 ◽  
Vol 32 (2) ◽  
pp. 255-275 ◽  
Author(s):  
R. E. Stephens
Keyword(s):  
Molecular Weight ◽  
Disulfide Bonds ◽  
Tertiary Structure ◽  
Sea Urchins ◽  
Guanidine Hydrochloride ◽  
Average Molecular Weight ◽  
Strongylocentrotus Droebachiensis ◽  
Optical Rotatory Dispersion ◽  
Partial Specific Volume ◽  
Mitotic Apparatus

The major 22S protein of the hexylene glycol-isolated mitotic apparatus has been characterized from spindle isolates and extracts of whole eggs and acetone powders of eggs from the sea urchins Strongylocentrotus purpuratus, Strongylocentrotus droebachiensis, and Arbacia punctulata. The protein is free of nucleotide, lipid, and ATPase activity. Essentially identical in amino acid composition, proteins from these species show a relatively high content of glutamic and aspartic acids and are fairly rich in hydrophobic amino acids. Optical rotatory dispersion studies indicate a helical content of about 20%, a value consistent with the proline content of the protein. The purified proteins have sedimentation rates in the range of 22–24S, diffusion constants of 2.4–2.5F, intrinsic viscosities of 3.7–4.3 ml/g, a partial specific volume of 0.74, and an average molecular weight of 880,000. Electron microscopy indicates a globular molecule with dimensions of approximately 150 by 200 A; such size and symmetry are consistent with hydrodynamic measurements. The 22S protein yields 6–7S, 9–10S, and 13–14S subunits below pH 4 or above pH 11. The 13–14S component has an estimated molecular weight of 600,000–700,000. A 5–6S particle is formed in 8 M urea or 5 M guanidine hydrochloride, while at pH 12 the 6–7S subunit is seen; each particle has a molecular weight of 230,000–240,000. In 8 M urea plus 2% mercaptoethanol or at pH 13, the molecular weight becomes 105,000–120,000; under these conditions the particle sediments at 2.5–3S and 4S, respectively. On the basis of these molecular weights, the 6–7S, 9–10S, 13–14S, and the parent 22S particle should be dimer, tetramer, hexamer, and octamer, respectively, of the 105,000–120,000 molecular weight subunit. The various subunits will reform the 22S particle when returned to neutral buffer, with the exception of the mercaptoethanol-treated urea subunit where breakage of disulfide bonds results in a polydisperse aggregate. The 22S particle itself is not susceptible to sulfhydryl reagents, implying either that the disulfide bonds are inaccessible or that they are unnecessary for maintenance of tertiary structure once the 22S particle has formed from subunits.

scholarly journals THE NATURE OF BENCE-JONES PROTEINS

1962 ◽  
Vol 116 (2) ◽  
pp. 207-227 ◽  
Author(s):  
G. M. Delman ◽  
J. A. Gally
Keyword(s):  
Molecular Weight ◽  
Extensive Study ◽  
Starch Gel Electrophoresis ◽  
Bence Jones Protein ◽  
Protein Amino Acid ◽  
Myeloma Protein ◽  
Γ Globulins ◽  
Normal Human ◽  
Polypeptide Chains ◽  
Bence Jones Proteins

The chemical relations among Bence-Jones proteins, myeloma proteins, and normal γ-globulins have been investigated by a variety of means. Starch gel electrophoresis in 8 M urea of reduced alkylated Bence-Jones proteins yielded patterns of bands corresponding to those of the light (L) polypeptide chains of the dissociated myeloma protein from the same patient. One instance in which this correspondence was found was chosen for extensive study. Chromatography on carboxymethylcellulose in 6 M urea was employed to isolate the light (L) polypeptide chains and heavy (H) polypeptide chains of the completely reduced and alkylated myeloma protein. Isolation of similarly treated Bence-Jones protein from the same patient corroborated the correspondence to the L chains of the myeloma protein. Amino acid analyses indicated that the compositions of the Bence-Jones protein and the L chains of the myeloma protein were identical. Moreover, the thermosolubility properties and spectrofluorometric behavior of the isolated L chains and Bence-Jones protein were similar. Ultracentrifugal analyses of the L chains of normal human 7S γ-globulin showed that their molecular weight in 6 M urea was 20,000. In aqueous solution their molecular weight was 41,000, suggesting that they exist as dimers under these conditions. The L chains of normal human γ-globulin were found to have reversible thermosolubility properties similar to those of Bence-Jones proteins. The H chains of normal human γ-globulin did not share these properties. Using spectrofluorometric methods, characteristic molecular transitions were found upon heating Bence-Jones proteins and L chains. These transitions were indicated by an increase in the intensity of fluorescence at well defined temperatures as well as by reversible shifts in the wavelength of maximal emission. The findings suggest that Bence-Jones proteins are composed of L chains of the type found in normal and pathological γ-globu]ins.

scholarly journals Development of Sample Pretreatment of Silk for Radiocarbon Dating

Radiocarbon ◽  
2008 ◽  
Vol 50 (1) ◽  
pp. 131-138 ◽  
Author(s):  
Kyeong Ja Kim ◽  
John Southon ◽  
Mineo Imamura ◽  
Rodger Sparks
Keyword(s):  
Molecular Weight ◽  
Radiocarbon Dating ◽  
Sample Pretreatment ◽  
Weight Fraction ◽  
Silk Protein ◽  
Age Dating ◽  
Different Types ◽  
Foreign Materials ◽  
Polypeptide Chains ◽  
Different Molecular Weight

We have developed sample pretreatments for silk for radiocarbon dating. Characteristics of silk under different types of pretreatment were investigated, as well as the behavior of dye and possible contaminants. We found that dye could be removed completely, together with all other foreign materials bigger than 1.2 μm, using a glass microfiber filter after decomposition with 6N HCl. The decomposed proteins were concentrated using Centriprep® ultrafiltration concentrators with 3 different molecular weight cut-offs. By taking a molecular weight fraction—which selects for secondary structures of silk protein—14C dating of silk samples can be made more reliable. This study confirms that uniformly fractured polypeptide chains of silk provide an appropriate fraction for 14C age dating to select silk protein against dye particles and undecomposed foreign contaminants.

Prospects for therapeutic use of fibronectin preparations

1986 ◽  
Vol 67 (5) ◽  
pp. 391-397
Author(s):  
R. I. Litvinov
Keyword(s):  
Molecular Weight ◽  
Extracellular Matrix ◽  
Endothelial Cells ◽  
Connective Tissue ◽  
Blood Plasma ◽  
Disulfide Bonds ◽  
Chemical Structure ◽  
Plasma Fibronectin ◽  
Adult Plasma ◽  
Polypeptide Chains

Blood plasma fibronectin is a glycoprotein with a molecular weight of 450 kD consisting of two almost identical polypeptide chains linked by disulfide bonds. The normal concentration of fibronectin in adult plasma is 0.3-0.4 g/l. Plasma fibronectin is synthesized by hepatocytes and possibly by endothelial cells. In its chemical structure and immunological properties it is similar to tissue fibronectin, which is formed mainly by connective tissue cells and is a part of extracellular matrix and collagen fibers.

The Structure of Radiation Cross-Linked Poly (ethylene oxide) Gels

1971 ◽  
Vol 29 ◽  
pp. 76-77
Author(s):  
C. E. Cluthe ◽  
G. G. Cocks
Keyword(s):  
Molecular Weight ◽  
Ethylene Oxide ◽  
Parallel Plate ◽  
Average Molecular Weight ◽  
Radical Reaction ◽  
Free Radical Reaction ◽  
Nitrogen Gas ◽  
Poly Ethylene Oxide ◽  
Poly Ethylene ◽  
Initial Viscosity

Aqueous solutions of a 1 weight-per cent poly (ethylene oxide) (PEO) were degassed under vacuum, transferred to a parallel plate viscometer under a nitrogen gas blanket, and exposed to Co60 gamma radiation. The Co60 source was rated at 4000 curies, and the dose ratewas 3.8x105 rads/hr. The poly (ethylene oxide) employed in the irradiations had an initial viscosity average molecular weight of 2.1 x 106.The solutions were gelled by a free radical reaction with dosages ranging from 5x104 rads to 4.8x106 rads.

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