Experimental nephritis due to type specific streptococci. VI. Further characterization of type 12 nephrotoxin

1969 ◽  
Vol 15 (6) ◽  
pp. 555-561 ◽  
Author(s):  
P. W. Fardy ◽  
B. H. Matheson ◽  
R. W. Reed
Keyword(s):  
Active Material ◽  
Gel Filtration ◽  
Acute Glomerulonephritis ◽  
Acid Hydrolysate ◽  
Culture Filtrates ◽  
Experimental Nephritis ◽  
Chemical Studies ◽  
Group A ◽  
High Voltage Electrophoresis

Additional studies were undertaken on the nature of a nephrotoxic agent found in the culture filtrates of certain group A streptococci. A commercially available dehydrated medium proved satisfactory for the production of the active material. Gel filtration was used to divide polypeptide extracts, prepared from the dialyzable portion of culture filtrates, into two major fractions. One of these, representing the higher molecular weight components, contained most of the nephrotoxic activity as evidenced by the development of hypertension and acute glomerulonephritis in rabbits injected with this fraction.Physical and chemical studies indicated that the active fraction consisted of at least four polypeptide components separable by high voltage electrophoresis on paper. Automatic amino acid analysis of an acid hydrolysate of this fraction revealed 17 different amino acids. Carbohydrate was not detected by anthrone and orcinol tests.No relationship was established between this streptococcal nephrotoxic agent and other streptococcal constituents which have been implicated in acute glomerulonephritis.

Purification and Characterization of Two Proteins from Culture Filtrates of Mycobacterium tuberculosis H 37 Ra Strain

1970 ◽  
Vol 1 (2) ◽  
pp. 164-168
Author(s):  
Thomas M. Daniel ◽  
Lavenia E. Ferguson
Keyword(s):  
Molecular Weight ◽  
Mycobacterium Tuberculosis ◽  
Skin Testing ◽  
Gel Filtration ◽  
Deae Cellulose ◽  
Purification And Characterization ◽  
Culture Filtrates ◽  
Zone Electrophoresis

Two proteins have been purified from culture filtrates of Mycobacterium tuberculosis , H 37 Ra strain by a procedure combining gel filtration, diethylaminoethyl (DEAE)-cellulose chromatography, and zone electrophoresis. The two proteins are similar in molecular weight but differ slightly in charge. The faster migrating protein, designated a 1 , is not antigenic. The slower migrating protein, designated a 2 , is antigenic both with respect to antisera and as a skin-testing antigen.

Elastase inhibitors in sputum from bronchitic patients with and without α1-proteinase inhibitor deficiency: partial characterization of a hitherto unquantified inhibitor of neutrophil elastase

1987 ◽  
Vol 73 (1) ◽  
pp. 19-28 ◽  
Author(s):  
H. M. Morrison ◽  
J. A. Kramps ◽  
S. C. Afford ◽  
D. Burnett ◽  
J. H. Dijkman ◽  
...  
Keyword(s):  
Neutrophil Elastase ◽  
Proteinase Inhibitor ◽  
Gel Filtration ◽  
Prolonged Incubation ◽  
Α2 Macroglobulin ◽  
Group A ◽  
Hydrolysis Of ◽  
Molecular Weight Fractions ◽  
Deficient Group

1. Anti-elastase function in sputum sol-phase from patients with α1-proteinase inhibitor (α1PI) deficiency was compared with sol-phase from patients with cigarette smoke-induced bronchitis and emphysema. 2. Both α1PI (2P < 0.01) and anti-leucoprotease (ALP) (2P < 0.01) concentrations were lower in sol-phase from the α1PI-deficient group, although α2-macroglobulin (α2M) levels were similar. 3. There was no difference in α1PI function between the two groups, but the inhibitor was only ≃ 30% active. 4. The absolute neutrophil elastase (NE) inhibitory capacity was similar in both groups (median 185 μg of NE inhibited/ml of sputum, range 80–480, for the α1PI-deficient group; median 175, range 80–300, for the bronchitic group). A substantial proportion of NE inhibition in secretions could not be accounted for by the amount of α1PI, ALP and α2M present (median 74.8%, range 43.2–97.4, for α1PI-deficient sol-phase; median 50.0%, range 0–80.8, for bronchitic sol-phase). 5. Gel filtration of sol-phase demonstrated the presence of NE inhibition in the low molecular weight fractions which was markedly sensitive to changes in substrate concentration and ionic strength, in contrast to purified α1PI and ALP. 6. Sputum sol-phase from both groups failed to prevent hydrolysis of elastin–fluorescein or succinyltrialanyl-p-nitroanilide by NE completely during prolonged incubation in the presence of an excess of functional inhibitors. This was more apparent in secretions from subjects with α1PI deficiency and may explain why such patients have a more rapidly progressive form of emphysema.

scholarly journals Innate Immunity in Lobsters: Partial Purification and Characterization of a Panulirus cygnus Anti-A Lectin

2012 ◽  
Vol 2012 ◽  
pp. 1-5 ◽  
Author(s):  
Robert L. P. Flower
Keyword(s):  
Innate Immunity ◽  
Polyacrylamide Gel Electrophoresis ◽  
Spiny Lobster ◽  
Gel Filtration ◽  
Partial Purification ◽  
Purification And Characterization ◽  
Rat Erythrocytes ◽  
Group A ◽  
Panulirus Cygnus

A lectin detected in haemolymph from the Australian spiny lobster Panulirus cygnus agglutinated human ABO Group A cells to a higher titre than Group O or B. The lectin also agglutinated rat and sheep erythrocytes, with reactivity with rat erythrocytes strongly enhanced by treatment with the proteolytic enzyme papain, an observation consistent with reactivity via a glycolipid. The lectin, purified by affinity chromatography on fixed rat-erythrocyte stroma, was inhibited equally by N-acetylglucosamine and N-acetylgalactosamine. Comparison of data from gel filtration of haemolymph (behaving as a 1,800,000 Da macromolecule), and polyacrylamide gel electrophoresis of purified lectin (a single 67,000 Da band), suggested that in haemolymph the lecin was a multimer. The purified anti-A lectin autoprecipitated unless the storage solution contained chaotropic inhibitors (125 mmol/L sucrose: 500 mmol/L urea). The properties of this anti-A lectin and other similar lectins are consistent with a role in innate immunity in these invertebrates.

scholarly journals Difference in the structural features of streptococcal M proteins from nephritogenic and rheumatogenic serotypes.

1987 ◽  
Vol 166 (1) ◽  
pp. 151-162 ◽  
Author(s):  
K M Khandke ◽  
T Fairwell ◽  
B N Manjula
Keyword(s):  
Tertiary Structure ◽  
Structural Information ◽  
Gel Filtration ◽  
Structural Features ◽  
Biologically Active ◽  
M Protein ◽  
Acute Glomerulonephritis ◽  
Group A Streptococci ◽  
M Proteins ◽  
Group A

The association of only certain M protein serotypes of group A streptococci with acute glomerulonephritis is very well recognized. Structural information on the M protein, a dimeric alpha-helical coiled-coil molecule, has come so far from three rheumatogenic serotypes, 5, 6, and 24. However, M proteins from the nephritogenic serotypes have not been well characterized. In the present study, we have isolated a biologically active 20,000 Mr pepsin fragment of type 49 M protein (PepM49), a nephritogenic serotype, and purified it to homogeneity using DEAE-Sephadex and gel filtration. The amino acid composition of PepM49 is similar to those of the rheumatogenic M protein serotypes PepM5, PepM6, and PepM24. However, the sequence of the NH2-terminal 60 residues of PepM49 shows little homology to any of these M protein serotypes, although the latter have significant homology among themselves. Nevertheless, PepM49 exhibits a strong heptad periodicity in its nonpolar residues, suggesting its overall conformational similarity with the other M molecules. During the course of the present studies, Moravek et al. (17) reported the NH2-terminal sequence of another M protein serotype, PepM1, which also does not exhibit much homology with the PepM5, PepM6, and PepM24 proteins. Our analysis of this sequence revealed that the PepM1 protein also exhibits a heptad periodicity of the nonpolar amino acids. A closer examination has revealed that the pattern of heptad periodicity in PepM49 and PepM1 proteins is more regular and more similar to each other than has been previously seen for the PepM5, PepM6, and PepM24 proteins. PepM1 is also a nephritogenic serotype. Taken together, these findings indicate an underlying conservation of the tertiary structure of the various M protein serotypes, despite the complexity in their antigenic variation and suggest that the nephritogenic M protein serotypes M1 and M49 may be further apart evolutionarily from the rheumatogenic serotypes 5, 6, and 24. The distinct differences in the structural features of the PepM1 and PepM49 proteins relative to the PepM5, PepM6, and PepM24 proteins are also suggestive of a correlation with the earlier broader classification of the group A streptococci into rheumatogenic and nephritogenic serotypes.

Characterization of an immunogenic fraction of Pasteurella multocida culture filtrates

1977 ◽  
Vol 23 (2) ◽  
pp. 197-201 ◽  
Author(s):  
Kunwar K. Srivastava ◽  
John W. Foster
Keyword(s):  
Culture Filtrate ◽  
Pasteurella Multocida ◽  
Gel Filtration ◽  
The Other ◽  
Rabbit Skin ◽  
Culture Filtrates ◽  
Homologous Challenge

An immunogenic fraction (IF) of Pasteurella multocida strain P-1059 was separated from culture filtrate by Sephadex gel filtration. Additional fractionation of IF with aqueous ether resulted in the glycoprotein-like preparation (GLP) while extraction with aqueous phenol provided the lipopolysaccharide-like preparation (LPP). The unextracted IF contained carbohydrate, protein, and lipid; the GLP contained carbohydrate and protein; and the LPP contained carbohydrate and lipid. The GLP was maximally protective for mice against homologous challenge, and was medially toxic in rabbit skin when compared to the other culture-filtrate preparations; the LPP was maximally toxic in rabbit skin, and was least protective for mice; and the unextracted IF was medially protective for mice, and was least toxic in rabbit skin.

Characterization of bacteriocin 28 produced by Clostridium perfringens

1982 ◽  
Vol 28 (7) ◽  
pp. 860-873 ◽  
Author(s):  
A. W. Li ◽  
J. A. Verpoorte ◽  
R. G. Lewis ◽  
D. E. Mahony
Keyword(s):  
Clostridium Perfringens ◽  
Density Gradient Centrifugation ◽  
Gradient Centrifugation ◽  
Polyacrylamide Gel Electrophoresis ◽  
Active Material ◽  
Sodium Dodecyl ◽  
Gel Filtration ◽  
Hydrophobic Properties ◽  
Isoelectric Points

Bacteriocin 28, produced by Clostridium perfringens, was characterized by gel filtration and sodium dodecyl sulfate – polyacrylamide gel electrophoresis as a glycoprotein with a molecular weight of approximately 100 000. Density gradient centrifugation suggested a lower weight of 84 000. The bacteriocin bound firmly to phenyl-Sepharose CL-4B gel, indicating hydrophobic properties, and elution from this gel with ethylene glycol clearly separated bacteriocin from the alpha and theta toxins of C. perfringens, the latter of which was also hydrophobic. Bacteriocin 28 was immunogenic, inducing neutralizing and precipitating antibodies, and possessed three isoelectric points: 7.37, 7.05, and 5.4. Amino acid and carbohydrate analysis of the active material showed a composition of 15 amino acids and several carbohydrates. The molecule demonstrated instability with increasing purification, and several approaches to purification are described.

scholarly journals Isolation, Purify and Characterization of Lectin from the Seeds of Artocarpus Species

2021 ◽  
pp. 10-15
Author(s):  
S. Krupa ◽  
K. R. Siddalinga Murthy
Keyword(s):  
Ion Exchange ◽  
Blood Group ◽  
Gel Filtration ◽  
Blood Groups ◽  
Exchange Chromatography ◽  
Buffer System ◽  
Hemagglutination Activity ◽  
Blood Group A ◽  
Group A

Aim: To isolate, partially purify and characterize lectin from the seeds of Artocarpus  species - using ammonium sulphate, gel-filtration and ion exchange chromatography.  Methodology: ABO blood groups were screened for the assay of hemagglutinating property with seed lectins of both. heterophyllus and A. hirsutus. The seed lectins were extracted using suitable buffer system pH 7.0 and partially purified by gel filtration and ion exchange chromatographic techniques. Results and Discussion: Lectins were extracted from the defatted seed powders of A. heterophyllus and A. hirsutus. The extracts were used for optimization of assay buffer for hemagglutination activity and 50 mM Tris-HCl buffer, pH 8.8 containing 1 mM CaCl2 and 1 mM MnCl2 showed greater hemagglutinating property. Screening for optimum concentration of RBCs from different blood group (A, B, AB and O groups) indicated O group at 10 % concentration as ideal for the assay of hemagglutinating property with seed lectins of both A. heterophyllus and A. hirsutus. Conclusion: In this study, lectin from the seeds of Artocarpus heterophyllus and Artocarpus hirsutus were isolated and partially purified. The isolated lectins were characterized for their heamagglutination activity. Among the human blood types (A+, B+, AB+ and O+) used, all the blood groups showed agglutination while greater agglutination was observed with O+ blood group.

Isolation and Characterization of Plasminogen Activator from Pig Leucocytes

1974 ◽  
Vol 31 (01) ◽  
pp. 072-085 ◽  
Author(s):  
M Kopitar ◽  
M Stegnar ◽  
B Accetto ◽  
D Lebez
Keyword(s):  
Molecular Weight ◽  
Plasminogen Activator ◽  
Gel Electrophoresis ◽  
Gel Filtration ◽  
Ph Optimum ◽  
Isolation And Characterization ◽  
Ph Range ◽  
Inhibition Studies ◽  
Final Purification

SummaryPlasminogen activator was isolated from disrupted pig leucocytes by the aid of DEAE chromatography, gel filtration on Sephadex G-100 and final purification on CM cellulose, or by preparative gel electrophoresis.Isolated plasminogen activator corresponds No. 3 band of the starting sample of leucocyte cells (that is composed from 10 gel electrophoretic bands).pH optimum was found to be in pH range 8.0–8.5 and the highest pH stability is between pH range 5.0–8.0.Inhibition studies of isolated plasminogen activator were performed with EACA, AMCHA, PAMBA and Trasylol, using Anson and Astrup method. By Astrup method 100% inhibition was found with EACA and Trasylol and 30% with AMCHA. PAMBA gave 60% inhibition already at concentration 10–3 M/ml. Molecular weight of plasminogen activator was determined by gel filtration on Sephadex G-100. The value obtained from 4 different samples was found to be 28000–30500.

The Immunological Characterization of Human Antibodies to Factor VIII Isolated by Immuno-Affinity Chromatography

1981 ◽  
Vol 45 (01) ◽  
pp. 060-064 ◽  
Author(s):  
M L Kavanagh ◽  
C N Wood ◽  
J F Davidson
Keyword(s):  
Factor Viii ◽  
Affinity Chromatography ◽  
Gel Filtration ◽  
Immunological Characterization ◽  
Human Antibodies ◽  
Heavy Chains ◽  
Light Chain Type ◽  
Antibody Preparations ◽  
Chain Type

SummaryNine human antibodies to factor VIII were isolated from haemophilic plasmas by affinity chromatography and gel filtration and six were subsequently subjected to immunological characterization. Three partially purified preparations were similarly characterized. Eight of the antibodies were characterized as being exclusively IgG and one preparation was found to contain IgM. Seven of the antibodies contained only a single light chain type, four being of type lambda and three of type kappa. Two antibody preparations contained both kappa and lambda light chains. In four of the preparations, only a single heavy chain sub-class could be demonstrated, three of IgG3 and one of IgG4. Of the remainder, three were a mixture of IgG3 and IgG4 sub-classes and one contained both IgG2 and IgG4. IgG sub-classification could not be achieved with the IgM-containing preparation. These results demonstrate a restricted heterogeneity of light and heavy chains in human antibodies to factor VIII.

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