Crambe Thioglucoside Glucohydrolase (EC 3.2.3.1): Separation of a Protein Required for Epithiobutane Formation

1973 ◽  
Vol 51 (12) ◽  
pp. 1654-1660 ◽  
Author(s):  
H. L. Tookey
Keyword(s):  
Ammonium Sulfate ◽  
Ferrous Ion ◽  
Seed Meal ◽  
Proteinaceous Material ◽  
Saturated Ammonium Sulfate ◽  
Crambe Seed

Sonicated extract of crambe seed meal prepared in the presence of ferrous ion and dithiothreitol enzymatically converts epi-progoitrin to glucose, HSO4−, and a mixture of 1-cyano-2-hydroxy-3,4-epithiobutanes (50–70%) and 1-cyano-2-hydroxy-3-butene (30–50%). A fraction of the extract precipitating between 60 and 70% saturated ammonium sulfate contains thioglucosidase that converts epi-progoitrin essentially to 1-cyano-2-hydroxy-3-butene. Chromatography (on cross-linked dextran) of a 40–60% ammonium sulfate fraction leads to separation of a proteinaceous material (s20 = 2.6 S) that does not hydrolyze epi-progoitrin but, in the presence of thioglucosidase, promotes the formation of 1-cyano-2-hydroxy-3,4-epithiobutanes in amounts proportional to those from crude seed meal extract.

Effect of organic reducing agents and ferrous ion on thioglucosidase activity of Crambe abyssinica seed

1970 ◽  
Vol 48 (9) ◽  
pp. 1024-1028 ◽  
Author(s):  
H. L. Tookey ◽  
I. A. Wolff
Keyword(s):  
Reducing Agents ◽  
Ferrous Ion ◽  
Seed Meal ◽  
Crambe Abyssinica ◽  
Meal Pattern ◽  
Crambe Seed ◽  
Crambe Meal ◽  
Insoluble Particles

The addition of L-ascorbate or 2-mercaptoethanol to aged crambe seed meal tends to restore the fresh meal pattern of epi-progoitrin hydrolysis to nitriles instead of (R)-goitrin. Neither of these reducing agents has an effect on the breakdown of epi-progoitrin to goitrin by an insoluble particulate thioglucosidase from crambe meal. The addition of ferrous ion to the insoluble particles results in the conversion of epi-progoitrin to (2S)-1-cyano-2-hydroxy-3-butene instead of (R)-goitrin over a range from pH 3.9 to 6.7.

MECHANISMS OF PLATELET AGGREGATION BY ACIDIC MUCOPOLYSACCHARIDE EXTRACTED FROM STICHOPUS JAPONICUS SELENKA IN HUMANS AND RABBITS

1987 ◽  
Author(s):  
J Z Li ◽  
E C-Y Lian
Keyword(s):  
Platelet Aggregation ◽  
Ammonium Sulfate ◽  
Human Platelets ◽  
Platelet Inhibitors ◽  
Rabbit Plasma ◽  
Human Fibrinogen ◽  
Stichopus Japonicus ◽  
Saturated Ammonium Sulfate ◽  
Rabbit Platelets ◽  
Induced Aggregation

It has been reported that acidic mucopolysaccharide extracted from sea cucumber (Stichopus japonicus selenka) (SJAMP) induced the aggregation of human and animal platelets by an unknown mechanism, using platelet-rich plasma (prp) and washed human and rabbit platelets we studied the effects of storage, platelet inhibitors, and various plasmas and their fractions on SJAMP-induced platelet aggregation. we found that the lowest concentrations of SJAMP required for aggregation of human and rabbit platelets were 0.4 and 2 ug/ml respectively. The reactivity of human platelets to SJAMP decreased with time after drawing of blood; rabbit platelets did not show this phenomenon. Platelet inhibitors such as aspirin, indomethacin, apyrase, antimycin, 2-deoxy-D-glucose, and EDTA inhibited by 50 to 100% the aggregation of human platelets induced by SJAMP; but these inhibitors had no effect on SJAMP-induced aggregation of rabbit platelets. Washed human and rabbit platelets were not aggregated by SJAMP. The aggregation of washed human platelets by SJAMP was restored completely by human or rabbit plasma, by human fibrinogen, or by 0 to 30% saturated ammonium sulfate fraction but not by serum. The aggregation of rabbit platelets by SJAMP could only be restored by rabbit plasma or serum, or by 50 to 60% saturated ammonium sulfate fraction. The data indicate that the mechanisms of aggregation of human and rabbit platelets by SJAMP are different. THe SJAMP-induced human platelet aggregation is dependent upon metabolism, release of ADP and the cyclooxygenase pathway requiring fibrinogen and Ca++. The aggregation of rabbit platelets induced by SJAMP is independent of metabolism, release of ADP and cyclooxygenase pathway, and does not require fibrinogen and Ca++, but needs certain protein(s) in the 50 to 60% saturated ammonium sulfate fraction of rabbit plasma.

scholarly journals CRYSTALLINE PNEUMOCOCCUS ANTIBODY

1949 ◽  
Vol 32 (6) ◽  
pp. 705-724 ◽  
Author(s):  
John H. Northrop ◽  
Walther F. Goebel
Keyword(s):  
Ammonium Sulfate ◽  
Horse Serum ◽  
Insoluble Fraction ◽  
Rabbit Serum ◽  
Type I ◽  
Type Ii ◽  
Serum Globulin ◽  
Saturated Ammonium Sulfate ◽  
Diphtheria Antitoxin ◽  
Specific Polysaccharide

1. The immune precipitate formed by antipneumococcus horse serum and the specific polysaccharide is not hydrolyzed by trypsin as is the diphtheria toxin-antitoxin complex, and purified pneumococcus antibody cannot be isolated by the method used for the isolation and crystallization of diphtheria antitoxin. 2. Type I pneumococcus antibody, completely precipitable by Type I polysaccharide, may be obtained from immune horse serum globulin by precipitation of the inert proteins with acid potassium phthalate. 3. The antibody obtained in this way may be fractionated by precipitation with ammonium sulfate into three main parts. One is insoluble in neutral salts but soluble from pH 4.5 to 3.0 and from pH 9.5 to 10.5. This is the largest fraction. A second fraction is soluble in 0.05 to 0.2 saturated ammonium sulfate and the third fraction is soluble in 0.2 saturated ammonium sulfate and precipitated by 0.35 saturated ammonium sulfate. The second fraction can be further separated by precipitation with 0.17 saturated ammonium sulfate to yield a small amount of protein which is soluble in 0.17 saturated ammonium sulfate but insoluble in 0.25 saturated ammonium sulfate. This fraction crystallizes in poorly formed, rounded rosettes. 4. The crystallization does not improve the purity of the antibody and is accompanied by the formation of an insoluble protein as in the case of diphtheria antitoxin. 5. None of the fractions obtained is even approximately homogeneous as determined by solubility measurements. 6. Purified antibody has also been obtained by dissociating the antigen-antibody complex. 7. The protective value of the fractions is quite different; that of the dissociated antibody being the highest and that of the insoluble fraction, the lowest. 8. All the fractions are immunologically specific since they do not precipitate with Type II polysaccharide nor protect against Type II pneumococci. 9. All the fractions give a positive precipitin reaction with antihorse rabbit serum. The dissociated antibody gives the least reaction. 10. Comparison of the various fractions, either by their solubility in salt solution or through immunological reactions, indicates that there are a large number of proteins present in immune horse serum, all of which precipitate with the specific polysaccharide but which have very different protective values, different reactions with antihorse rabbit serum, and different solubility in salt solutions.

Diastereomeric episulfides from epi-progoitrin upon autolysis of crambe seed meal

1968 ◽  
Vol 7 (6) ◽  
pp. 989-996 ◽  
Author(s):  
M.E. Daxenbichler ◽  
C.H. VanEtten ◽  
I.A. Wolff
Keyword(s):  
Seed Meal ◽  
Crambe Seed

Solubilization and Selected Properties of Crambe Seed Thioglucosidase (Thioglucoside Glucohydrolase, EC 3.2.3.1)

1973 ◽  
Vol 51 (9) ◽  
pp. 1305-1310 ◽  
Author(s):  
H. L. Tookey
Keyword(s):  
Substrate Inhibition ◽  
Apparent Molecular Weight ◽  
Ferrous Ion ◽  
Soluble Enzyme ◽  
Ammonium Sulfate Precipitation ◽  
Michaelis Constant ◽  
Sequential Fractionation ◽  
Crambe Abyssinica ◽  
Crambe Seed ◽  
Insoluble Particles

Ultrasonic treatment releases thioglucoside glucohydrolase (thioglucosidase) from insoluble particles of Crambe abyssinica seed meal. The crude enzyme is optimally activated by 10−2 M ascorbate, requires a reducing agent for stability, and is inhibited by 10−3 M p-chloromercuribenzenesulfonate.Chromatography of a meal extract on cross-linked dextran separates two enzyme fractions. About 80% of the activity elutes at an apparent molecular weight of 110 000; the remainder elutes at the void volume. Both crude soluble enzyme and the fractions from the dextran column produce goitrin from epi-progoitrin, but in the presence of ferrous ion the chief aglucon product becomes 1-cyano-2-hydroxy-3-butene. The Michaelis constant (of the major peak) at pH 5 is 0.004 M. Ferrous ion produces substrate inhibition at high epi-progoitrin levels. Sequential fractionation of meal extract by ammonium sulfate precipitation and by chromatography on cross-linked dextran effects an 80-fold purification.

Selektive Anreicherung von Staphylokokken-Panton-Valentine-Leukozidin mit Aluminiumoxid-Cholesterin / Selective Enrichment of Staphylococcus “Panton-Valentine”-Leukocidine with Aluminium Oxide Cholesterol

1969 ◽  
Vol 24 (11) ◽  
pp. 1431-1435 ◽  
Author(s):  
M. Kanoe ◽  
H. Blobel ◽  
W. Schaeg
Keyword(s):  
Staphylococcus Aureus ◽  
Ammonium Sulfate ◽  
Culture Supernatant ◽  
Aluminium Oxide ◽  
Sulfate Solution ◽  
Ammonium Sulfate Solution ◽  
Pressure Filtration ◽  
Selective Enrichment ◽  
Saturated Ammonium Sulfate

A selective concentration of “Panton-Valentine”-leukozidine (PVL) from the culture-supernatant Staphylococcus aureus was achieved through adsorption of most of the a-hemolysin, coagulase, eggyolk-opacity factor and fibrinolysin to aluminiumoxide cholesterol. PVL was further concentrated through dialysis against a saturated ammonium sulfate solution and subsequent pressure-filtration. Through pressure-filtration relatively low-molecular staphylococcal substances, such as a part of a nuclease, could be separated from PVL.

scholarly journals THE CRYSTALLIZATION AND SEROLOGICAL DIFFERENTIATION OF A STREPTOCOCCAL PROTEINASE AND ITS PRECURSOR

1950 ◽  
Vol 92 (3) ◽  
pp. 201-218 ◽  
Author(s):  
S. D. Elliott
Keyword(s):  
Ammonium Sulfate ◽  
Crystalline Form ◽  
Antigenic Specificity ◽  
Serological Tests ◽  
Saturated Ammonium Sulfate ◽  
Alkaline Reaction ◽  
Group A ◽  
Refrigerator Temperature ◽  
Streptococcal Proteinase ◽  
Degree Of Purification

Grown in dialysate broth at a pH between 5.5 and 6.5, some strains of group A streptococci elaborate the precursor of a proteolytic enzyme. Within this range of hydrogen concentration the precursor is also produced when the streptococci are suspended in a peptone dialysate containing glucose and incubated at 37°C. The precursor does not appear to be produced at a neutral or alkaline reaction. Methods are described whereby the precursor and proteinase have been isolated in crystalline form. The precursor crystallizes from half-saturated ammonium sulfate at pH 8.0 and a temperature of 22°C. or higher; the proteinase crystallizes from 0.15 saturated ammonium sulfate at pH 8.0 but does so most readily at refrigerator temperature. The degree of purification achieved by these procedures is discussed. The activity of purified preparations of the precursor and of proteinase has been tested against α-benzoyl-l-arginineamide and, with this as a substrate, the conversion of precursor to proteinase by autocatalysis or by trypsin has been confirmed. Immunological experiments are described, the results of which provide evidence of the distinct antigenic specificity of the precursor and proteinase; the conversion of precursor to proteinase has been followed by means of serological tests.

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