scholarly journals The molecular weight of J chains derived from human immunoglobulin M

1974 ◽  
Vol 137 (1) ◽  
pp. 71-77 ◽  
Author(s):  
Manuel J. Ricardo ◽  
John M. Brewer ◽  
Franklin P. Inman
Keyword(s):  
Molecular Weight ◽  
Saline Solution ◽  
Guanidine Hydrochloride ◽  
Average Molecular Weight ◽  
Sodium Dodecyl ◽  
Immunoglobulin M ◽  
Human Immunoglobulin ◽  
Average Weight ◽  
Polyacrylamide Gels ◽  
J Chain

J chain was isolated from sulphonated human immunoglobulin M molecules by electrophoresis on polyacrylamide gels. When determined by electrophoresis in sodium dodecyl sulphate–polyacrylamide gels, the molecular weight of the protein was about 27000. After suspension in 5m-guanidine hydrochloride solution for 21 days, two groups of three bands appeared on the gels. Most of the protein dissociated to components of molecular weight 15000. The molecular weight of purified J chain was also determined by ultracentrifugation. In borate–saline solution the average weight-average molecular weight was about 29000. The molecular weight slowly decreased upon prolonged exposure to guanidine hydrochloride, and after 14 days the minimum molecular weight was about 15000. Some association between chains still existed. These data suggest that J chain derived from the paraprotein exists in borate–saline solution as dimers held by strong non-covalent forces.

scholarly journals The incorporation of J chain into reassembled human immunoglobulin M

1974 ◽  
Vol 137 (1) ◽  
pp. 79-83 ◽  
Author(s):  
Manuel J. Ricardo ◽  
Franklin P. Inman
Keyword(s):  
Molecular Weight ◽  
High Molecular Weight ◽  
Immunoglobulin M ◽  
Human Immunoglobulin ◽  
Fc Fragment ◽  
Guanidinium Hydrochloride ◽  
J Chain ◽  
Disulphide Bonds ◽  
Complete Dissociation ◽  
Schlieren Pattern

Human IgM (immunoglobulin M) was reduced with 24mm-mercaptoethylamine. This atreatment resulted in complete dissociation to IgMs subunits and free J chain. Intr-subunit interchain disulphide bonds remained intact. The mixture then was encouraged to reoxidize. The schlieren pattern of the reoxidized mixture showed the presence of a considerable quantity of IgM in addition to residual IgMs. The isolated reassembled IgM did not dissociate in 5m-guanidinium hydrochloride. It apparently contained the same amount of covalently attached J chain as did native IgM. The J chain was a part of the high-molecular-weight Fc fragment obtained from the reassembled IgM.

scholarly journals Investigations of the structural function of the J chain in human immunoglobulin M

1973 ◽  
Vol 131 (4) ◽  
pp. 677-682 ◽  
Author(s):  
Manuel J. Ricardo ◽  
Franklin P. Inman
Keyword(s):  
Gel Filtration ◽  
Elution Curve ◽  
Immunoglobulin M ◽  
Human Immunoglobulin ◽  
Structural Function ◽  
J Chain ◽  
Mild Reduction

Human IgM molecules were treated with Na2SO3 or mercaptoethylamine in concentrations ranging from 2 to 14mm or 2 to 22mm respectively. The dissociation of IgM to IgMs varied from 0% to 100%. At the intermediate concentrations of either reagent the amount of freed J chains was less than expected. In an attempt to find an explanation for this, IgM was partially dissociated to IgMs with mercaptoethylamine. The IgMs isolated by gel filtration was divided according to the ascending and descending portions of the elution curve. These portions were treated with 24mm-mercaptoethylamine and analysed for the presence of J chains. Only the ascending portion contained free J chains. Thus, after mild reduction where not all the IgM molecules are dissociated to IgMs, some J chains remain covalently attached to some IgMs molecules although most of the J chains are freed. It was concluded that the J chain could serve as a ‘hitch’ for IgMs molecules forming intact IgM.

Calcium-binding proteins in a vorticellid contractile organelle

1975 ◽  
Vol 19 (1) ◽  
pp. 203-213
Author(s):  
W.B. Amos ◽  
L.M. Routledge ◽  
F.F. Yew
Keyword(s):  
Amino Acids ◽  
Molecular Weight ◽  
Calcium Binding ◽  
Binding Proteins ◽  
Sodium Dodecyl ◽  
Aromatic Amino Acids ◽  
Calcium Binding Proteins ◽  
Polyacrylamide Gels ◽  
Protein Species ◽  
Low Concentrations

The proteins of the contractile spasmoneme of Zoothamnium have been examined for comparison with other motile systems. Though capable of calcium-induced contraction, glycerinated preparations of the spasmoneme contain neither actin nor tubulin at levels that can be detected in polyacrylamide gels. Sixty per cent of the protein in sodium dodecyl sulphate gels migrates in a band at a molecular weight of approximately 20,000, consisting largely of 2 similar protein species which are here given the name of spasmins. The amino acid composition of 2 spasmin fractions has been determined by a fluorimetric method. They are rich in Asx, Glx and serine, but have few aromatic amino acids and no cystine or methionine. In calcium-buffered polyacrylamide gels, it was observed that a reduction in the electrophoretic mobility of the spasmins was induced specifically by calcium (but not magnesium) at the same low concentrations as induce contraction. This indicates that the spasmins are calcium-binding proteins which may be involved directly in the calcium-induced contraction of the spasmoneme.

scholarly journals Transformation-deficient mutants of Bacillus subtilis impaired in competence-specific nuclease activities

1982 ◽  
Vol 152 (1) ◽  
pp. 166-174
Author(s):  
J A Mulder ◽  
G Venema
Keyword(s):  
Molecular Weight ◽  
Bacillus Subtilis ◽  
Sodium Dodecyl Sulfate ◽  
Sodium Dodecyl ◽  
Magnesium Ions ◽  
Polyacrylamide Gels ◽  
Wild Type ◽  
Molecular Weights ◽  
Defective Mutant ◽  
Mutant Strains

A comparison of the nucleolytic activities in competent and physiologically low-competent wild-type cultures of Bacillus subtilis in DNA-containing sodium dodecyl sulfate-polyacrylamide gels revealed the existence of three competence-associated nuclease activities with apparent molecular weights of 13,000, 15,000, and 26,000. The three activities, which were dependent on manganese or magnesium ions, were specifically present in the competent fraction of a competent culture. The competence-associated nucleolytic activities of eight transformation-defective mutant strains were assayed, resulting in the following three classes of mutants: (i) four strains which, according to this assay, were not impaired in any of the nucleolytic activities mentioned above; (ii) one strain which was strongly impaired in the 13,000- and 26,000-molecular-weight activities, but showed a considerable level of the 15,000-molecular-weight activity; and (iii) three strains which were severely impaired in all three activities. The results indicated that the 26,000-molecular-weight activity was a dimer of the 13,000-molecular-weight activity and that this nuclease was involved in the entry of DNA.

The isolation of surface array proteins from bacteria

1984 ◽  
Vol 62 (11) ◽  
pp. 1181-1189 ◽  
Author(s):  
S. F. Koval ◽  
R. G. E. Murray
Keyword(s):  
Molecular Weight ◽  
Protein Conformation ◽  
Gel Electrophoresis ◽  
Guanidine Hydrochloride ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Structural Features ◽  
Proteolytic Degradation ◽  
Low Concentrations ◽  
Single Polypeptide

The methods used for the isolation of regularly structured (RS) surface array proteins of a range of prokaryotes are described. Most RS proteins can be selectively solubilized from envelope preparations with low concentrations of urea or guanidine hydrochloride. Sodium dodecyl sulfate – polyacrylamide gel electrophoresis analysis of the protein extracts shows that most RS arrays are composed of a single polypeptide that may contain carbohydrate. The molecular weight of the proteins varies from 41 000 to 200 000. Possible reasons for the presence of more than one polypeptide in RS protein preparations are discussed, as well as the evidence for proteolytic degradation of some RS proteins during isolation. Structural features of the RS proteins are described and the importance of protein conformation to assembly of the arrays is indicated.

scholarly journals ISOLATION AND REACTIVATION OF THE AXOSTYLE

1973 ◽  
Vol 56 (1) ◽  
pp. 13-26 ◽  
Author(s):  
Mark S. Mooseker ◽  
Lewis G. Tilney
Keyword(s):  
Molecular Weight ◽  
Sodium Dodecyl Sulfate ◽  
Atpase Activity ◽  
Adenosine Triphosphate ◽  
Sodium Dodecyl ◽  
Polyacrylamide Gels ◽  
Molecular Weights ◽  
Dodecyl Sulfate ◽  
Cilia And Flagella ◽  
Triton X

The contractile axostyle is a ribbon-shaped organelle present in certain species of flagellates found in the hindgut of wood eating insects. This organelle propagates an undulatory wave whose motion, like flagella and cilia, is related to microtubules. Unlike the axoneme of cilia and flagella, however, the axostyle is composed of singlet microtubules linked together in parallel rows. Axostyles were isolated from Cryptocercus gut protozoa with Triton X-100. Normal motility of the isolated axostyle could be restored with adenosine triphosphate (ATP); the specific conditions necessary for this reactivation were essentially identical with those reported for the reactivation of isolated flagella or whole sperm. ATPase activity of the isolated axostyle was comparable to the values reported for ciliary or flagellar axonemes. The axostyle was reasonably specific for ATP. Most of the proteins of the isolated axostyle comigrated with proteins of the ciliary axoneme on sodium dodecyl sulfate (SDS) polyacrylamide gels (i e. equivalent molecular weights). These included the following: the higher molecular weight component of dynein, tubulin, linkage protein (nexin), and various secondary proteins. Evidence for dynein in the axostyle is presented and a model proposed to explain how repeated propagated waves can be generated.

scholarly journals Detection in milk of a 60,000 Mr mouse and a 68,000 Mr human phosphorylated glycoprotein by histochemical analysis on polyacrylamide gels.

1983 ◽  
Vol 31 (6) ◽  
pp. 709-716 ◽  
Author(s):  
M R Green
Keyword(s):  
Molecular Weight ◽  
Sialic Acid ◽  
Periodic Acid ◽  
Sodium Dodecyl ◽  
Apparent Molecular Weight ◽  
Blue Staining ◽  
Polyacrylamide Gels ◽  
Periodic Acid Schiff ◽  
Coomassie Blue Staining ◽  
Coomassie Blue

Proteins in colostrum and skimmed milk from humans and mice were separated by electrophoresis on polyacrylamide gels and stained with Coomassie blue (CB), Ethyl-Stains-all (ESA), and periodic acid-Schiff (PAS) to investigate changes that may occur in milks throughout lactation. In mouse colostrum but not in mature mouse milk, a PAS-positive protein of apparent molecular weight of 60,000 stained prominently blue with ESA. A protein in human milk with a molecular weight of 68,000 stained similarly but was present throughout lactation. The intensity of blue staining of these minor proteins in milk approached that obtained with casein phosphoproteins. The metachromatic dye ESA stains phosphoproteins and sialic acid-rich glycoproteins blue to blue-green. Removal of phosphorus from the former and sialic acid from the latter results in those proteins staining red with ESA. The intensity of blue staining of the 60,000 and 68,000 Mr proteins was diminished but not lost following treatment with phosphatase. It was eliminated following neuraminidase digestion of the mouse protein and mild acid hydrolysis of the human protein. Coomassie blue staining of the proteins was not affected by these procedures. Following electrophoresis of milk and milk fractions in a non-sodium dodecyl sulfate-containing system, the proteins were identified by their characteristic staining properties with ESA and isolated.

Isolation and Characterization of J-chain from Bovine Colostral Immunoglobulin M

1975 ◽  
Vol 53 (9) ◽  
pp. 943-949 ◽  
Author(s):  
R. Komar ◽  
T. K. S. Mukkur
Keyword(s):  
Gel Electrophoresis ◽  
Fast Moving ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Polyacrylamide Gel ◽  
Immunoglobulin M ◽  
J Chain ◽  
Isolation And Characterization ◽  
Moving Band ◽  
Covalently Bound

Purified bovine colostral intact immunoglobulin M (IgM) exhibited the presence of an anodal, single, fast moving band (noncovalently bound form) when subjected to analytical polyacrylamide gel electrophoresis at an alkaline pH in urea. Reduced and alkylated or sulfitolysed bovine colostral IgM (devoid of the noncovalently bound form) also showed the presence of a similar band (covalently bound form). The molecular weight of both the covalently bound and noncovalently bound forms of the fast component was determined to be 16 500 by sodium dodecyl sulfate – polyacrylamide gel electrophoresis. In addition, the non-covalently bound form of the fast-moving component was found to be antigenically identical to the covalently bound form. The noncovalently bound form sedimented as a single peak at 1.56 S. Antiserum against the fast-moving component precipitated neither bovine colostral IgG nor μ-chains and bovine serum albumin, but precipitated native or denatured intact IgM (devoid of the non-covalently bound form) and human J-chains and vice versa, thus permitting the fast-moving components to be classified as J-chains. Radioalkylation experiments revealed the presence of 9.7 sulfhydryl groups per mole, for both the covalently and non-covalently bound forms of bovine J-chain. The stoichiometry of J-chain, determined from the densitometric tracing of the reduced and alkylated bovine colostral IgM (devoid of the noncovalently bound J-chain) in stained analytical polyacrylamide gels, revealed the presence of one J-chain per IgM molecule. On the other hand the amount of non-covalently bound form of J-chain was determined to be 1.2 per molecule of IgM.

scholarly journals Lectin-binding proteins in central-nervous-system myelin. Detection of glycoproteins of purified myelin on polyacrylamide gels by [3h]concanavalin A binding

1979 ◽  
Vol 183 (2) ◽  
pp. 205-212 ◽  
Author(s):  
L J McIntyre ◽  
R H Quarles ◽  
R O Brady
Keyword(s):  
Molecular Weight ◽  
Rat Brain ◽  
Concanavalin A ◽  
Binding Proteins ◽  
Lectin Binding ◽  
Sodium Dodecyl ◽  
Myelin Proteins ◽  
Polyacrylamide Gels ◽  
Myelin Associated Glycoprotein

Concanavalin A strongly agglutinates purified fragments of immature and mature rat brain myelin, but only weakly agglutinates mature bovine and human myelin fragments. A sensitive method involving [3H]concanavalin binding to sodium dodecyl sulphate/polyacrylamide gels was used to detect the concanavalin A-binding proteins in purified myelin. When applied to mature rat brain myelin proteins that had been labelled in vivo with [14C]fucose, the distribution of the [3H]concanavalin A on the gel was very similar to that of [14C]fucose with the major peak corresponding to the major myelin-associated glycoprotein. The technique revealed that the immature form of the myelin-associated glycoprotein with a slightly larger apparent molecular weight also bound concanavalin A, and that in purified immature rat myelin the quantitative importance of some of the other glycoproteins in binding concanavalin A was increased relative to the myelin-associated glycoprotein. The separated proteins of bovine and human myelin bound more [3H]-concanavalin A than those of rat myelin. In these species, the myelin-associated glycoprotein was a major concanavalin A-binding protein, although two higher-molecular-weight glycoproteins also bound significant quantities of [3H]concanavalin A. The results indicate that there are receptors for concanavalin A on the surface of rat, bovine and human myelin membranes and suggest that the myelin-associated glycoprotein is one of the principal receptors.

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