THE PURIFICATION OF BEEF THYROGLOBULIN WITH THE USE OF GEL FILTRATION

1963 ◽  
Vol 41 (12) ◽  
pp. 2493-2500 ◽  
Author(s):  
L. Perelmutter ◽  
W. Devlin ◽  
N. R. Stephenson
Keyword(s):  
Thyroid Gland ◽  
Serum Proteins ◽  
Narrow Band ◽  
Protein A ◽  
Gel Filtration ◽  
Thyroid Extract ◽  
Saline Extract ◽  
Sedimentation Constant ◽  
Minced Beef ◽  
Fraction I

A method is described for preparing a 19 S thyroglobulin component from a saline beef thyroid extract by chromatographic fractionation on Sephadex G-200. By employing Sephadex G-200 it was possible to separate a saline extract of minced beef thyroid gland into several fractions, the first of which (fraction I) contained approximately 60% of the protein and 96% of the iodine. Spectrophotometric, ultracentrifugal, and immunochemical methods revealed that fraction I possessed, in addition to the 19 S thyroglobulin protein, a 25 S component as well as serum proteins. A narrow band of fraction I appeared to be free of both the 25 S component and the serum proteins. The material in this subfraction had a sedimentation constant of S20,w = 19.1 and contained 1.2 mg of iodine per 100 mg of protein.

THE PURIFICATION OF BEEF THYROGLOBULIN WITH THE USE OF GEL FILTRATION

1963 ◽  
Vol 41 (1) ◽  
pp. 2493-2500 ◽  
Author(s):  
L. Perelmutter ◽  
W. Devlin ◽  
N. R. Stephenson
Keyword(s):  
Thyroid Gland ◽  
Serum Proteins ◽  
Narrow Band ◽  
Protein A ◽  
Gel Filtration ◽  
Thyroid Extract ◽  
Saline Extract ◽  
Sedimentation Constant ◽  
Minced Beef ◽  
Fraction I

A method is described for preparing a 19 S thyroglobulin component from a saline beef thyroid extract by chromatographic fractionation on Sephadex G-200. By employing Sephadex G-200 it was possible to separate a saline extract of minced beef thyroid gland into several fractions, the first of which (fraction I) contained approximately 60% of the protein and 96% of the iodine. Spectrophotometric, ultracentrifugal, and immunochemical methods revealed that fraction I possessed, in addition to the 19 S thyroglobulin protein, a 25 S component as well as serum proteins. A narrow band of fraction I appeared to be free of both the 25 S component and the serum proteins. The material in this subfraction had a sedimentation constant of S20,w = 19.1 and contained 1.2 mg of iodine per 100 mg of protein.

scholarly journals Purification and partial characterization of four proteins from human parotid saliva

1971 ◽  
Vol 123 (3) ◽  
pp. 455-464 ◽  
Author(s):  
Anders Bennick ◽  
George E. Connell
Keyword(s):  
Serum Proteins ◽  
Protein A ◽  
Gel Filtration ◽  
Binding Activity ◽  
Significant Activity ◽  
Parotid Saliva ◽  
Polyacrylamide Electrophoresis ◽  
Ph Range ◽  
Human Parotid Saliva

Four proteins, which have been designated A, B, C and D, have been purified from human parotid saliva. These proteins are the major constituents of parotid saliva which migrate rapidly to the anode in polyacrylamide electrophoresis at pH9.5. Gel filtration and polyacrylamide electrophoresis were employed in the purification procedures. After purification all four preparations were tested for homogeneity by electrophoresis at pH2.8 and 9.5, by isoelectric focusing in the pH range 3–10, by immunodiffusion, and by sedimentation in the analytical ultracentrifuge. None of the proteins showed significant activity in assays for amylase, acid and alkaline phosphatase, protease, lysozyme, ribonuclease, peroxidase, β-glucuronidase, β-galactosidase, iron-binding activity and esterase. No cross-reactions were detected with antisera specific for lactoferrin and 15 serum proteins. All four proteins were rich in glutamic acid, proline and glycine and were lacking completely the sulphur-containing amino acids. Proteins A and C contained no threonine or tyrosine. Carbohydrate could be demonstrated only in protein A at a concentration of 4% of the total protein.

EPIDEMIC THYROIDITIS

1964 ◽  
Vol 45 (3) ◽  
pp. 381-401 ◽  
Author(s):  
G. Hintze ◽  
P. Fortelius ◽  
J. Railo
Keyword(s):  
Thyroid Gland ◽  
Distribution Pattern ◽  
Needle Biopsy ◽  
Serum Proteins ◽  
Acute Infection ◽  
Thyroid Tissue ◽  
Subacute Thyroiditis ◽  
Present Observation ◽  
Upper Respiratory Tract ◽  
Endemic Goitre

ABSTRACT A type of subacute thyroiditis occurring epidemically in a factory in Helsinki was observed in 44 cases. In every case the thyroiditis followed an acute infection of the upper respiratory tract. The variation in incidence during one and a half years was in good agreement with that of the acute infection. Since Helsinki is in an endemic goitre region, the fact that the disease was of the migrating type was of great diagnostic importance. In all cases but one, the nodules have persisted. One case of asymptomatic thyroiditis was seen. In the majority of the patients the thyroid gland had been carefully palpated before the thyroiditis occurred, and in all cases the condition was followed up by the same investigator. Special attention was paid to changes in the iodine metabolism, the serum cholesterol, the electrophoretic distribution pattern of the serum proteins, and the circulating thyroid auto-antibodies. In many cases needle biopsy of the thyroid gland was performed. Thyroid function invariably returned to normal with time, although one patient remained in a hypothyroid state for about a year. In no cases were thyroid auto-antibodies found. For the beta-globulin fraction, the electrophoretic distribution pattern of the serum proteins gave values which were still not normalized in any case, and only in two cases was the alpha2-fraction normalized. The needle biopsy, when thyroid tissue was obtained, showed almost the same picture as in endemic goitre, but in some specimens nonspecific inflammatory changes were seen. Prednisolone relieved the symptoms, but did not affect the course of the disease. According to the present observation this type of epidemic thyroiditis would seem to represent a form of nonspecific subacute thyroiditis.

scholarly journals Surfactant protein D binding to alveolar macrophages

1994 ◽  
Vol 300 (1) ◽  
pp. 237-242 ◽  
Author(s):  
K Miyamura ◽  
L E A Leigh ◽  
J Lu ◽  
J Hopkin ◽  
A López Bernal ◽  
...  
Keyword(s):  
Alveolar Macrophages ◽  
Serum Proteins ◽  
Specific Binding ◽  
Lectin Binding ◽  
Protein A ◽  
Direct Interaction ◽  
Surfactant Protein ◽  
Lung Epithelial Cells ◽  
Surfactant Protein D ◽  
Apparent Dissociation Constant

Surfactant protein D (SP-D) is a lung-specific protein, synthesized and secreted by lung epithelial cells. It belongs to group III of the family of C-type lectins; each member of this group has an unusual overall structure consisting of multiple globular ‘head’ regions (which contain the C-type lectin domains) linked by triple-helical, collagen-like, strands. This group includes the surfactant protein A (SP-A) and the serum proteins mannan-binding protein, conglutinin and collectin-43, all of which have been shown to bind to the C1q receptor found on a wide variety of cells, including macrophages. Both SP-D and SP-A have been shown to enhance oxygen radical production by alveolar macrophages. Although this strongly suggests a direct interaction between SP-D and a specific receptor on alveolar macrophages, it is still unclear whether SP-D binds to the same receptor used by SP-A and/or C1q. Human SP-D was isolated from amniotic fluid and was radiolabelled using 125I. Alveolar macrophages were isolated from human bronchioalveolar lavage fluid, and also from bovine lung washings, by differential adhesion to 24-well tissue-culture plates. The study was carried out using EDTA-containing buffers, to eliminate Ca(2+)-dependent C-type lectin binding, and was also carried out at 4 degrees C to eliminate possible internalization by the cells. 125I-SP-D showed specific binding to alveolar macrophages in both a time- and concentration-saturable manner. The binding was inhibited, by approx. 90%, on addition of a 200-fold excess of unlabelled SP-D. The apparent dissociation constant (Kd) was (3.6 +/- 1.3) x 10(-11) M, based on the assumption that native SP-D is assembled as a dodecamer of 12 identical polypeptides of 43 kDa to yield a protein of 516 kDa. C1q was also shown to bind alveolar macrophages (Kd 3 x 10(-6) M), but addition of C1q did not show inhibition of the binding of 125I-SP-D to the macrophages. We conclude that SP-D binds specifically to alveolar macrophages and the receptor involved is different from that utilized by C1q.

scholarly journals Human serum protein fractionation by gel filtration

1970 ◽  
Vol 118 (5) ◽  
pp. 869-873 ◽  
Author(s):  
T. Freeman ◽  
J. Smith
Keyword(s):  
Human Serum ◽  
Serum Protein ◽  
Serum Proteins ◽  
Gel Filtration ◽  
Protein Fractionation ◽  
Logical Approach ◽  
Complex Protein ◽  
Human Serum Protein ◽  
Immunological Technique ◽  
Current Terminology

The development of a quantitative immunological technique using polyvalent antiserum permits a more logical approach to the fractionation of complex protein mixtures. In this study whole serum was separated by conventional gel filtration and the fractions obtained were analysed. This demonstrates over 60 immunologically distinct serum proteins. Because the current terminology is inadequate to describe this number of proteins, a temporary numerical nomenclature has been used.

scholarly journals Presence in human eosinophils of a lysophospholipase similar to that found in the pancreas

1995 ◽  
Vol 309 (1) ◽  
pp. 141-144 ◽  
Author(s):  
F W Holtsberg ◽  
L E Ozgur ◽  
D E Garsetti ◽  
J Myers ◽  
R W Egan ◽  
...  
Keyword(s):  
Synthetic Peptide ◽  
Protein A ◽  
Pancreatic Enzyme ◽  
Gel Filtration ◽  
Pancreatic Tissue ◽  
Supernatant Fraction ◽  
Polyadenylated Rna ◽  
Rabbit Polyclonal Antibody ◽  
Dna Fragment ◽  
Lysophospholipase Activity

The supernatant fraction from lysed human eosinophils, when separated by gel-filtration chromatography, contains a protein with lysophospholipase activity of approximate molecular mass 74 kDa. This mass differs substantially from the 17 kDa of a previously cloned eosinophil lysophospholipase (Charcot-Leyden crystal protein), but is similar to that reported for a pancreatic enzyme. We have therefore further characterized this pancreatic-like lysophospholipase in human eosinophils. A rabbit polyclonal antibody was produced against a synthetic peptide consisting of amino acids 325-349 from the 74 kDa rat pancreatic lysophospholipase. Western-blot analysis of eosinophil extracts indicate that this antibody recognizes a single 74 kDa band in these preparations. Incubation of the supernatant fraction from sonified eosinophils with this antibody, followed by precipitation of antibody-antigen complexes with Protein A, removes the majority of the lysophospholipase activity. Indirect immunofluorescence examination with this antibody indicates this protein to be localized to granules of eosinophils and not in other leucocytes. Moreover, reverse transcriptase PCR of polyadenylated RNA from eosinophils and from rat pancreatic tissue with primers to rat pancreatic lysophospholipase resulted in readily detectable 1 kb DNA products in both samples. Sequencing revealed this DNA fragment to be identical with the human pancreatic lysophospholipase cDNA sequence. Taken together, these data indicate that eosinophils contain a lysophospholipase that is similar to the human pancreatic enzyme.

scholarly journals THE OCCURRENCE DURING ACUTE INFECTIONS OF A PROTEIN NOT NORMALLY PRESENT IN THE BLOOD

1947 ◽  
Vol 85 (5) ◽  
pp. 491-498 ◽  
Author(s):  
Maclyn McCarty
Keyword(s):  
Infectious Diseases ◽  
Early Phase ◽  
Serum Proteins ◽  
Protein A ◽  
Normal Serum ◽  
C Reactive Protein ◽  
Reactive Protein ◽  
Acute Infections

A procedure is described for the isolation and crystallization from human serous fluids of the C-reactive protein, a substance which appears in the blood especially in the early phase of certain acute infectious diseases. Immunological studies confirm earlier work in showing that the protein is highly antigenic and serologically specific, and demonstrate that crystallization of the protein effectively separates it from normal serum proteins.

scholarly journals Human diadenosine triphosphate hydrolase: preliminary characterisation and comparison with the Fhit protein, a human tumour suppressor.

2000 ◽  
Vol 47 (2) ◽  
pp. 435-441 ◽  
Author(s):  
A C Asensio ◽  
C R Rodríguez-Ferrer ◽  
S Oaknin ◽  
P Rotllán
Keyword(s):  
Protein A ◽  
Gel Filtration ◽  
Human Platelets ◽  
Tumour Suppressor ◽  
Human Tumour ◽  
Fhit Protein ◽  
Diadenosine Triphosphate

Human platelets diadenosine triphosphatase was characterised and compared with the Fhit protein, a human tumour suppressor with diadenosine triphosphatase activity. Both enzymes exhibit similar Km, are similarly activated by Mg2+, Ca2+ and Mn2+, and inhibited by Zn2+ and suramin. However, they are differentially inhibited by Fhit antibodies and exhibit differences in gel-filtration behaviour.

scholarly journals On the identity of vitronectin and S-protein: immunological crossreactivity and functional studies

Blood ◽  
1986 ◽  
Vol 68 (3) ◽  
pp. 737-742
Author(s):  
BR Tomasini ◽  
DF Mosher
Keyword(s):  
Monoclonal Antibody ◽  
Serum Proteins ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Gel Filtration ◽  
Membrane Attack Complex ◽  
Biochemical Properties ◽  
Complex Data ◽  
Heparin Binding ◽  
S Protein

Vitronectin (serum spreading factor), a major serum cell adhesion molecule, was compared with S-protein, the inhibitor of the C5–9 membrane attack complex. Data from the literature indicate that S- protein and vitronectin are alpha globulins with the same aminoterminal residues, amino acid compositions, and concentrations in normal plasma (150 to 250 micrograms/mL). Both proteins have been reported to interact with the thrombin-antithrombin complex. The cDNA sequences of vitronectin and S-protein were recently determined and found to be almost identical. In the present studies, rabbit-anti-S-protein and a monoclonal antibody to vitronectin both recognized 65,000- and 75,000- molecular weight (mol wt) polypeptides when plasma or serum proteins were separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and transferred to nitrocellulose paper. The 65,000 and 75,000-mol wt polypeptides bound more avidly from serum than plasma to monoclonal anti-vitronectin or heparin coupled to agarose. The presence or absence of the polypeptides constituted a major difference between the heparin-binding proteins of serum and plasma. When complement- activated serum and unactivated serum were separated by gel filtration, vitronectin coeluted with C9 in high-mol-wt fractions of activated serum but not unactivated serum. Purified S-protein was recognized by the monoclonal antibody to vitronectin and promoted spreading of human skin fibroblasts. Both vitronectin and S-protein were degraded by thrombin. On the basis of immunological and functional, as well as biochemical, properties, therefore, S-protein and vitronectin are the same.

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