The Peroxidase of Dioscorea esculenta: Partial Purification and Characterisation

1999 ◽  
Vol 54 (7-8) ◽  
pp. 496-500 ◽  
Author(s):  
J. Okpuzor ◽  
O. Omidiji
Keyword(s):  
Molecular Weight ◽  
Ion Exchange ◽  
Neutral Type ◽  
Gel Filtration ◽  
Apparent Molecular Weight ◽  
Partial Purification ◽  
Optimum Temperature ◽  
Total Peroxidase Activity ◽  
Lower Size ◽  
Parent Activation

Abstract Dioscorea esculenta. Peroxidase. Ion Exchange Chrom atography. Gel Filtration We isolated two types of peroxidase from the fresh tuber of Dioscorea esculenta using a combination of ion exchange and gel filtration chrom atography. The results showed one type to be neutral and the other to be strongly ionic. The strongly ionic type constitutes 70% of total peroxidase activity in the tissue. The apparent molecular weight of the neutral type is 38 kD a while the anionic type has an apparent molecular weight of 57 kD a. It was possible with the use of gel filtration on Sephadex G-200 followed by FPLC on phenyl superose to purify the lower size POD by a factor of 15, while the larger ionic peroxidase was purified 68 fold com pared to the crude with protein yields of 0.90% and 1.30% respectively. The ionic POD is m ore therm o-stable, has a higher optimum temperature for activity and has a higher ap parent activation energy com pared to the neutral POD from this source.

Characterisation of a Neutral Protease Produced by Micrococcus luteus

1996 ◽  
Vol 51 (5-6) ◽  
pp. 429-431 ◽  
Author(s):  
M.O. Ilori ◽  
O.O. Amund ◽  
O. Omidiji
Keyword(s):  
Molecular Weight ◽  
Citric Acid ◽  
Proteolytic Enzyme ◽  
Micrococcus Luteus ◽  
Gel Filtration ◽  
Apparent Molecular Weight ◽  
Ion Exchange Chromatography ◽  
Exchange Chromatography ◽  
Optimum Temperature ◽  
Optimum Ph

Abstract A proteolytic enzyme produced by a cassava-ferment­ing strain of Micrococcus luteus was extracted and puri­fied 50-fold by gel filtration and ion exchange chromatography. The optimum pH for the enzyme was 7.0, the optimum temperature 25 °C, the apparent molecular weight 42 kDa and the Km value, 0.45 mg ml-1 with casein as substrate. The enzyme was stimulated by Ca2+ and Mg2+ but inhibited by Zn2+ and Co2+ ions. Other inhibitors were EDTA, KCN, citric acid and L-cysteine indicating the enzyme to be a metalloprotease.

Purification and properties of an exocellular β-glucosidase of Candida molischiana (Zikes) Meyer and Yarrow capable of hydrolyzing soluble cellodextrins

1985 ◽  
Vol 63 (11) ◽  
pp. 1160-1166 ◽  
Author(s):  
Pierre Gondé ◽  
Robert Ratomahenina ◽  
Alain Arnaud ◽  
Pierre Galzy
Keyword(s):  
Molecular Weight ◽  
Ion Exchange ◽  
Gel Filtration ◽  
Ion Exchange Chromatography ◽  
Exchange Chromatography ◽  
Optimum Temperature ◽  
Purification And Properties ◽  
Optimum Ph

The exocellular enzyme β-glucosidase of Candida molischiana was studied. This strain is able to ferment soluble cellodextrins. The enzyme was partially purified by ion-exchange chromatography and gel filtration. The molecular weight of this enzyme was 120 000; its optimum pH was between 4 and 4.5 and its optimum temperature was 60 °C. This enzyme was active against different soluble glucosides and was inhibited by p-chloromercuribenzoate, gluconolactone, and glucose. A "glucosyltransferase" activity appeared in the presence of ethanol. The biosynthesis of the enzyme was constitutive but repressed by glucose.

scholarly journals Purification of the Acetyl CoACarboxylase-1 from Serum of Breast Cancer Women after Mastectomy

2021 ◽  
Vol 2 (4) ◽  
pp. 32-42
Author(s):  
Sabah Gazal ◽  
Susan Jamil Ali . ◽  
Perry Habib Saifullah .
Keyword(s):  
Breast Cancer ◽  
Molecular Weight ◽  
Ion Exchange ◽  
Substrate Concentration ◽  
Gel Filtration ◽  
Deae Cellulose ◽  
Single Band ◽  
Optimum Temperature ◽  
Menten Constant ◽  
Optimal Ph

Acetyle CoA Carboxylase-1was purified from serum of premenop- ausal women with breast cancer (after Mastectomy or treatment ) by Gel Filtration using Sephadex G-100 and by Ion Exchange using DEAE-Cellulose A-50, also the molecular weight was estimated by the Electrophoresis on Acrylamide in the absence of denaturing elements . The result showed that a single band was obtained at 210 KD by Gel Filtering while Ion Exchange showed one band at 210 KD. The optimum temperature of purified Acetyle CoA Carboxylase-1 was 40 , optimal pH at 7.5 and the optimum substrate concentration at 1.8mM. Michaelis-Menten constant (km) was 0.3mM and Vmax was 23mM.min-1.

Physico-Chemical Properties of Recombinant Desulphatohirudin

1990 ◽  
Vol 63 (03) ◽  
pp. 499-504 ◽  
Author(s):  
A Electricwala ◽  
L Irons ◽  
R Wait ◽  
R J G Carr ◽  
R J Ling ◽  
...  
Keyword(s):  
Molecular Weight ◽  
Gel Filtration ◽  
Chemical Properties ◽  
Alpha Helix ◽  
Apparent Molecular Weight ◽  
Correlation Spectroscopy ◽  
Nmr Studies ◽  
Recombinant Hirudin ◽  
Physico Chemical ◽  
Recombinant Molecule

SummaryPhysico-chemical properties of recombinant desulphatohirudin expressed in yeast (CIBA GEIGY code No. CGP 39393) were reinvestigated. As previously reported for natural hirudin, the recombinant molecule exhibited abnormal behaviour by gel filtration with an apparent molecular weight greater than that based on the primary structure. However, molecular weight estimation by SDS gel electrophoresis, FAB-mass spectrometry and Photon Correlation Spectroscopy were in agreement with the theoretical molecular weight, with little suggestion of dimer or aggregate formation. Circular dichroism studies of the recombinant molecule show similar spectra at different pH values but are markedly different from that reported by Konno et al. (13) for a natural hirudin-variant. Our CD studies indicate the presence of about 60% beta sheet and the absence of alpha helix in the secondary structure of recombinant hirudin, in agreement with the conformation determined by NMR studies (17)

Studies on an Inhibitor of Plasminogen Activation in Human Serum

1973 ◽  
Vol 30 (02) ◽  
pp. 414-424 ◽  
Author(s):  
Ulla Hedner
Keyword(s):  
Ion Exchange ◽  
Human Serum ◽  
Antithrombin Iii ◽  
Gel Filtration ◽  
Ion Exchange Chromatography ◽  
Exchange Chromatography ◽  
Specific Adsorption ◽  
Partial Purification ◽  
Plasminogen Activation ◽  
Preparative Electrophoresis

SummaryA procedure is described for partial purification of an inhibitor of the activation of plasminogen by urokinase and streptokinase. The method involves specific adsorption of contammants, ion-exchange chromatography on DEAE-Sephadex, gel filtration on Sephadex G-200 and preparative electrophoresis. The inhibitor fraction contained no antiplasmin, no plasminogen, no α1-antitrypsin, no antithrombin-III and was shown not to be α2 M or inter-α-inhibitor. It contained traces of prothrombin and cerulo-plasmin. An antiserum against the inhibitor fraction capable of neutralising the inhibitor in serum was raised in rabbits.

Partial purification of a boar sperm membrane protein inducing sperm agglutinating antibody

1990 ◽  
Vol 10 (2) ◽  
pp. 131-139
Author(s):  
Oyewole Adeyemo ◽  
E. O. Okegbile ◽  
O. O. Olorunsogo
Keyword(s):  
Molecular Weight ◽  
Membrane Protein ◽  
Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Gel Filtration ◽  
Partial Purification ◽  
Boar Sperm ◽  
Sperm Membrane ◽  
Triton X

For the development of immunological contraception, attention is being concentrated on the possibility of using a sperm membrane antigen. Boar sperm membrane was extracted with triton-X 100 and fractionated by Sephadex G-150 column chromatography. The glycosylated and nonglycosylated portions of protein peaks from the gel filtration were obtained by fractionating on concanavalin A-Sepharose and eluting the bound protein with 0.3 M methyl mannoside. A glycosylated fraction was found to induce sperm agglutinating antibodies in rabbit. The partially purified protein has a molecular weight of 30 kilodaltons, as determined by sodium dodecyl polyaccyrlamide gel electrophoresis. Further work is planned on the histochemical determination of the origin of this protein and species cross-activity of the antibody.

Acid proteases from species of Mucor. III. Interaction with concanavalin A and concanavalin A Sepharose

1976 ◽  
Vol 54 (2) ◽  
pp. 120-129 ◽  
Author(s):  
W. S. Rickert ◽  
P. A. McBride-Warren
Keyword(s):  
Molecular Weight ◽  
Concanavalin A ◽  
Gel Filtration ◽  
Apparent Molecular Weight ◽  
Affinity Column ◽  
Mucor Miehei ◽  
Highly Active ◽  
Elution Buffer ◽  
Turbidimetric Assay ◽  
Ph Range

The reaction of Mucor miehei protease with concanavalin A was followed by a turbidimetric assay in the pH range 5–8. At pH 4.0, no turbidity developed but binding of the enzyme to concanavalin A could be demonstrated by gel filtration. Two fractions of apparent molecular weight 65 000 and 52 000 were isolated, the 65 000 molecular weight species apparently representing a protomer of concanavalin A (24 000) bound to the enzyme. An analysis of the circular dichroism spectrum of this complex suggested that protomer binding results in a conformational change in the enzyme which is associated with a 30% increase in proteolytic activity.At pH 6.0, the enzyme was strongly bound to columns of concanavalin A Sepharose but could be removed by including α-methyl D-glucoside and NaCl in the elution buffer. Some column degradation occurred at room temperature but was not detectable at 4 °C where rapid elution of the enzyme resulted in a greater than 90% yield of highly active protein. Periodate-oxidized Mucor miehei protease and Mucor rennin did not react with concanavalin A and were not bound to the affinity column.

scholarly journals Effects of pronase and neuraminidase treatment on a myelin-associated glycoprotein in developing brain

1976 ◽  
Vol 156 (1) ◽  
pp. 143-150 ◽  
Author(s):  
R H Quarles
Keyword(s):  
Molecular Weight ◽  
Sialic Acid ◽  
Sodium Dodecyl Sulphate ◽  
Sodium Dodecyl ◽  
Gel Filtration ◽  
Apparent Molecular Weight ◽  
Weight Class ◽  
Developmental Difference ◽  
Adult Rats ◽  
Neuraminidase Treatment

Rats (14 days old) were injected with [14c]fucose and young adult rats with [3H]fucose in order to label the myelin-associated glycoproteins. As previously reported, the major [14C]fucose-labelled glycoprotein in the immature myelin had a higher apparent molecular weight on sodium dodecyl sulphate/polyacrylamide gels that the [3H]fucose-labelled glycoprotein in mature myelin. This predominant doubly labelled glycoprotein component was partially purified by preparative gel electrophoresis and converted to glycopeptides by extensive Pronase digestion. Gel filtration on Sephadex G-50 separated the glycopeptides into several clases, which were designted A,B, C AND D, from high to low molecular weight. The 14C-labelled glycopeptides from immature myeline were enriched in the highest-molecular-weight class A relative to the 3H-labelled glycopeptides from mature myelin. Neuraminidase treatment of the glycoprotein before Pronase digestion greatly decreased the proportion of glycopeptides fractionating in the higher-molecular-weight classes and largely eliminated the developmental differences that were apparent by gel filtration. However, neuraminidase treatment did not decrease the magnitude of the developmental difference revealed by electrophoresing the intact glycoprotein on sodium dodecyl sulphate gels, although it did decrease the apparent molecular weight of the glycoprotein from both the 15-day-old and adult rats by an amount comparable in magnitude to that developmental difference. The results from gel filtration of glycopeptides indicate that there is a higher content of large molecular weight, sialic acid-rich oligosaccharide units in the glycoprotein of immature myelin. However, the higher apparent molecular weight for the glycoprotein from 15-day-old rats on sodium dodcyl sulphate gels is not due primarily to its higher sialic acid content.

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.

Partial purification and properties of an L-arabinose dehydrogenase from Azospirillum brasiliense

1983 ◽  
Vol 29 (2) ◽  
pp. 242-246 ◽  
Author(s):  
Norman J. Novick ◽  
Max E. Tyler
Keyword(s):  
Molecular Weight ◽  
Divalent Cation ◽  
Gel Filtration ◽  
Reducing Agent ◽  
Partial Purification ◽  
Ph Optimum ◽  
Glycine Buffer ◽  
Purification And Properties ◽  
Filtration Technique

An L-arabino-aldose dehydrogenase responsible for the oxidation of L-arabinose to L-arabino-γ-lactone has been purified 59-fold from L-arabinose grown cells of Azospirillum brasiliense. The dehydrogenase was found to be specific for substrates with the L-arabino-configuration at carbons 2, 3, and 4. Km values for L-arabinose of 75 and 140 μM were found with NADP and NAD as coenzymes, respectively. The enzyme had a pH optimum of 9.5 in glycine buffer and was stable when heated to 55 °C for 5 min. No enhancement of activity in the presence of any divalent cation or reducing agent tested was found. L-Arabinose dehydrogenase had a molecular weight of 175 000 as measured by the gel filtration technique.

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