scholarly journals The purification and properties of ferritin from human serum

1976 ◽  
Vol 157 (1) ◽  
pp. 97-103 ◽  
Author(s):  
M Worwood ◽  
S Dawkins ◽  
M Wagstaff ◽  
A Jacobs
Keyword(s):  
Anion Exchange ◽  
Serum Ferritin ◽  
Serum Proteins ◽  
Polyacrylamide Gel Electrophoresis ◽  
Gel Filtration ◽  
Anion Exchange Chromatography ◽  
Exchange Chromatography ◽  
Partial Purification ◽  
Human Spleen ◽  
Human Ferritin

1. Ferritin has been isolated from the serum of four patients with iron overload by using two methods. 2. In method A, the serum was adjusted to pH 4.8 and heated to 70 degrees C. After removal of denatured protein, ferritin was concentrated and further purified by ion-exchange chromatography and gel filtration. In most cases, only a partial purification was achieved. 3. In method B, ferritin was extracted from the serum with a column of immuno-adsorbent [anti-(human ferritin)] and released from the column with 3M-KSCN. Further purification was achieved by anion-exchange chromatography followed by the removal of remaining contaminating serum proteins by means of a second immunoadsorbent. Purifications of up to 31 000-fold were achieved, and the homogeneity of the final preparations was demonstrated by polyacrylamide-gel electrophoresis. 4. Serum ferritin purified by either method has the same elution volume as human spleen ferritin on gel filtration on Sephadex G-200. Serum ferritin has a relatively low iron content and iron/protein ratios of 0.023 and 0.067 (mug of Fe/mug of protein) were found in two pure preparations. On anion-exchange chromatography serum ferritin has a low affinity for the column when compared with various tissue ferritins. Isoelectric focusing has demonstrated the presence of a high proportion of isoferritins of relatively high pI. 5. Possible mechanisms for the release of ferritin into the circulation are briefly discussed.

scholarly journals Purification and Characterization of Two Endo-β-1,4-glucanases from Mollusca, Ampullaria crossean

2005 ◽  
Vol 37 (10) ◽  
pp. 702-708 ◽  
Author(s):  
Yan-Hong Li ◽  
Rui Guo ◽  
Qiu-Yu Yin ◽  
Ming Ding ◽  
Si-Liang Zhang ◽  
...  
Keyword(s):  
Anion Exchange ◽  
Carboxymethyl Cellulose ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecylsulfate ◽  
Gel Filtration ◽  
Residual Activity ◽  
Hydrolytic Activity ◽  
Anion Exchange Chromatography ◽  
Exchange Chromatography ◽  
The Stability

Abstract Two novel endo-β-1,4-glucanases, EG45 and EG27, were isolated from the gastric juice of mollusca, Ampullaria crossean, by anion exchange, hydrophobic interaction, gel filtration and a second round of anion exchange chromatography. The purified proteins EG45 and EG27 appeared as a single band on sodium dodecylsulfate polyacrylamide gel electrophoresis with a molecular mass of 45 kDa and 27 kDa, respectively. The optimum pH for CMC activity was 5.5 for EG45 and 4.4-4.8 for EG27. The optimum temperature range for EG27 was broad, between 50 °C and 60 °C; for EG45 it was 50 °C. The analysis on the stability of these two endo-β-1,4-glucanases showed that EG27 was acceptably stable at pH 3.0-11.0 even when the incubation time was prolonged to 24 h at 30 °C, whereas EG45 remained relatively stable at pH 5.0-8.0. About 85% of the activity of EG27 could be retained upon incubation at 60 °C for 24 h. However, less than 10% residual activity of EG45 was detected at 50 °C. Among different kinds of substrates, both enzymes showed a high preference for carboxymethyl cellulose. EG45, in particular, showed a carboxymethyl cellulose hydrolytic activity of 146.5 IU/mg protein. Both enzymes showed low activities to xylan (from oat spelt) and Sigmacell 101, and they were inactive to p-nitrophenyl-β-D-cellobioside, salicin and starch.

Form and action of a protease in cotyledons of germinating peanut seeds

1972 ◽  
Vol 50 (11) ◽  
pp. 2189-2195 ◽  
Author(s):  
P. N. R. Mainguy ◽  
R. B. van Huystee ◽  
D. B. Hayden
Keyword(s):  
Anion Exchange ◽  
Polyacrylamide Gel Electrophoresis ◽  
Gel Filtration ◽  
Temperature Response ◽  
Anion Exchange Chromatography ◽  
Exchange Chromatography ◽  
Ammonium Sulfate Precipitation ◽  
Ph Optimum ◽  
Stokes Radius ◽  
Precipitation Ph

The extract from cotyledons of 6-day-old peanut seedlings contained an enzyme that decomposed the chromogenic substrate N-benzoyl-D,L-arginine-p-nitroanilide (BAPA). The enzyme was purified by ammonium sulfate precipitation, pH adjustment, and anion-exchange chromatography. The molecular weight was found to be 60 000 as estimated from the Stokes radius obtained by gel filtration on Sephadex G 200. Its temperature response curve showed optimal activity between 25 and 36° and its pH optimum was 7.4. Anion-exchange chromatography as well as polyacrylamide gel electrophoresis of the purified extract resulted in two distinct areas of activity with regard to BAPA hydrolysis. Inhibitor experiments revealed that the enzyme does not have sulfhydryl groups at its active site nor does it respond to specific trypsin inhibitors, but it was inactivated by diisopropylfluorophosphate. It is therefore likely that it is a serine-type peptidase.

scholarly journals Isolation, partial purification and characterization of phospholipase A2 from Naja Katiensis venom

2021 ◽  
Vol 13 (2) ◽  
pp. 107-112
Author(s):  
C.F. Okechukwu ◽  
P.L. Shamsudeen ◽  
R.K. Bala ◽  
B.G. Kurfi ◽  
A.M. Abdulazeez
Keyword(s):  
Ion Exchange ◽  
Phospholipase A2 ◽  
Specific Activity ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Gel Filtration ◽  
Ion Exchange Chromatography ◽  
Exchange Chromatography ◽  
Partial Purification ◽  
Crude Venom

The most effective and acceptable therapy for snakebite victims is the immediate administration of antivenin which is limited by problems of hypersensitivity reactions in some individuals and its inability to resolve the local effects of the venom. The aim of this study was to isolate, partially purify and characterize phospholipase A2 from Naja Katiensis venom. Phospholipase A2 was partially purified via a two-step process: gel filtration on Sephadex G-75 and ion exchange chromatography using CM Sephadex, and subjected to SDS-PAGE analysis. From the results, the specific activity of the partially purified PLA2 decreased from 0.67μmol/min/mg in crude venom to 0.29μmol/min/mg after ion exchange chromatography with a yield of 5% and purification fold of 0.43. The optimum temperature of the purified PLA2 was found to be 35ºC and optimum p.H of 7. velocity studies for the determination of kinetic constants using L-a-lecithin as substrate revealed a Km  of 1.47mg/ml and Vmax  of 3.32μ moles/min/mg. The sodium dodecyl sulphate polyacrylamide gel electrophoresis of the purified PLA2 showed a distinct band with molecular weight estimated to be 14KDa. In conclusion, the present study shows that phospholipase A2 was isolated, purified and characterized. This may serve as a promising candidate for future development of a novel anti-venin drug.

scholarly journals Interferon alpha associated with systemic lupus erythematosus is not intrinsically acid labile.

1989 ◽  
Vol 169 (3) ◽  
pp. 987-993 ◽  
Author(s):  
A M Yee ◽  
J P Buyon ◽  
Y K Yip
Keyword(s):  
Anion Exchange ◽  
Lupus Erythematosus ◽  
Molecular Size ◽  
Anion Exchange Chromatography ◽  
Exchange Chromatography ◽  
Partial Purification ◽  
Ifn Alpha ◽  
Plasma Factor ◽  
Acid Stable ◽  
Acid Labile

The physicochemical properties of apparently acid-labile IFN-alpha from patients with SLE have been studied. The antigenicity, apparent molecular size, and isoelectric point of SLE IFN-alpha are indistinguishable from those of conventional, previously characterized, acid-stable subspecies of IFN-alpha. However, after partial purification by anion-exchange chromatography, SLE IFN-alpha no longer exhibits acid lability, suggesting that other plasma factor(s) are responsible for the acid lability of SLE IFN-alpha. Addition of SLE plasma, but not normal plasma, to conventional acid-stable IFN-alpha renders the exogenous IFN-alpha acid labile. Preliminary results demonstrate that an acid-dependent IFN-inactivating activity can be partially purified from SLE plasma by anion-exchange chromatography.

Identification of Glucagon-Like Peptide-1(7–36) Amide in Rat Brain

1989 ◽  
Vol 26 (2) ◽  
pp. 169-171 ◽  
Author(s):  
S Yoshimoto ◽  
M Hirota ◽  
C Ohboshi ◽  
K Shima
Keyword(s):  
Rat Brain ◽  
Anion Exchange ◽  
Gel Filtration ◽  
Anion Exchange Chromatography ◽  
Exchange Chromatography ◽  
Experimental Conditions ◽  
Specific Radioimmunoassay ◽  
Glucagon Like Peptide ◽  
Glucagon Like Peptide 1 ◽  
The Brain

Acid-urea extract of rat brain was examined by glucagon-like peptide-1 (GLP-1) specific radioimmunoassay. A single peak was observed which co-eluted with GLP-1(7–36)amide on gel filtration and anion exchange chromatography. In contrast, GLP-1(1–37) was not detected under our experimental conditions. The fact that GLP-1 (7–36)amide, but not GLP-1(1–37), was present in rat brain suggests that preproglucagon was processed in the brain in the same manner as in the intestine and not as in the pancreas.

scholarly journals Mouse Uterine Alkaline Phosphatase: Improved Purification by Affinity Chromatography and Further Characterization of the Enzyme

1980 ◽  
Vol 33 (3) ◽  
pp. 279 ◽  
Author(s):  
RN Murdoch ◽  
Louise E Buxton ◽  
DJ Kay
Keyword(s):  
Alkaline Phosphatase ◽  
Affinity Chromatography ◽  
Polyacrylamide Gel Electrophoresis ◽  
Gel Filtration ◽  
Deae Cellulose ◽  
Apparent Molecular Weight ◽  
Anion Exchange Chromatography ◽  
Pregnant Mouse ◽  
Exchange Chromatography ◽  
Enzyme Preparations

An improved procedure for the purification of alkaline phosphatase from about 10 g of day 7 pregnant mouse uterine tissue is described. Following homogenization, the procedure involved solubilization and extraction with 0�8% (v/v) Triton X-lOO and 20% (v/v) n-butanol, ammonium sulfate precipitation, concanavalin A-Sepharose 4B affinity chromatography, DEAE-cellulose anion-exchange chromatography and Sephacryl S200 gel filtration. On subjecting 2162-fold purified enzyme preparations to polyacrylamide-gel electrophoresis, a single band of protein coincident with the zone of enzyme activity and having an apparent molecular weight of 205 OOO� lOOOO was identified. Affinity chromatography yielded the largest increase in purity of any step in the procedure and established the glycoprotein nature of the uterine enzyme.

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