Inorganic pyrophosphatase of Streptococcus faecium F24

1972 ◽  
Vol 18 (2) ◽  
pp. 183-192 ◽  
Author(s):  
Patricia R. Starr ◽  
Evelyn L. Oginsky
Keyword(s):  
Stationary Phase ◽  
Specific Activity ◽  
Deae Cellulose ◽  
Inorganic Pyrophosphatase ◽  
Inactivation Kinetics ◽  
Starch Gel Electrophoresis ◽  
Diethylaminoethyl Cellulose ◽  
Single Protein ◽  
Streptococcus Faecium ◽  
Specific Activities

The acid Co2+-activated and the alkaline Mg2+-activated inorganic pyrophosphatases of Streptococcus faecium F24 were studied by a variety of techniques to determine whether they are activities of two distinct proteins or of a single protein. Both enzyme activities were found to be cryptic and soluble. The specific activities of both enzymes increased coordinately 1.5- to 3-fold during exponential growth at 37C, and then decreased as the cells approached stationary phase. Similar shifts in specific activity did not occur upon growth at 30C. The specific activities in extracts of cells in stationary phase varied coordinately over a 2-fold range depending on nutritional conditions. Diethylaminoethyl cellulose (DEAE-cellulose) and Sephadex G-100 column chromatography and starch–gel electrophoresis did not resolve the two activities. Slight differences in the thermal inactivation kinetics of the two activities were observed. It was concluded that the two pyrophosphatase activities are most probably those of a single protein under different assay conditions.

scholarly journals The cell wall of Bacillus licheniformis N.C.T.C. 6346. Biosynthesis of the teichuronic acid

1966 ◽  
Vol 101 (3) ◽  
pp. 692-697 ◽  
Author(s):  
R.C. Hughes
Keyword(s):  
Stationary Phase ◽  
Bacillus Licheniformis ◽  
Specific Activity ◽  
Deae Cellulose ◽  
Paper Electrophoresis ◽  
Exponential Phase ◽  
Teichuronic Acid ◽  
High Concentrations ◽  
Flavobacterium Heparinum ◽  
Testicular Hyaluronidase

1. Particulate fractions prepared from disrupted cells of Bacillus licheniformis N.C.T.C. 6346 catalyse the uptake of radioactivity from UDP-[(14)C]glucuronic acid or UDP-N[(14)C]-acetylglucosamine. Maximal uptake requires the presence of both nucleotides and Mg(2+) ions. The reaction is inhibited markedly by high concentrations of novobiocin and, to a certain extent, by vancomycin and by methicillin. 2. The radioactive product formed is resistant to Pronase and is soluble in 5% (w/v) trichloroacetic acid. It is of high molecular weight, from its behaviour on columns of Sephadex G-50 or G-200, and behaves during paper electrophoresis in n-acetic acid and chromatography on DEAE-cellulose in a manner similar to teichuronic acid. 3. Both teichuronic acid and the synthesized material are resistant to testicular hyaluronidase and to Flavobacterium heparinum heparinase. 4. The specific activity of suspensions of broken cells or of washed particulate fractions is greatest when they are prepared from exponentially growing cells. Fractions obtained from late exponential-phase or stationary-phase cells have very low activity. 5. The galactosamine content of B. licheniformis N.C.T.C. 6346 cell walls increases during the exponential phase and decreases during the stationary phase.

Studies on the relation between plasma antithrombin and heparin-cofactor

1968 ◽  
Vol 46 (3) ◽  
pp. 347-350 ◽  
Author(s):  
F. C. Monkhouse ◽  
Susan Milojevic
Keyword(s):  
Gel Electrophoresis ◽  
Specific Activity ◽  
Deae Cellulose ◽  
Fold Increase ◽  
Significant Loss ◽  
Starch Gel Electrophoresis ◽  
Electrophoretic Patterns ◽  
Starch Gel ◽  
Cofactor Activity ◽  
Cellulose Column

A method for the preparation of purified plasma antithrombin and heparin-cofactor is described. The method involves adsorption by aluminium hydroxide, separation on a DEAE-cellulose column by means of a graded salt concentration, and vertical curtain electrophoresis. A 100-fold increase in the specific activity of antithrombin and a 30-fold increase in the specific activity of heparin-cofactor have been achieved. In spite of the increased purification, no separation of the two activities was achieved. When a highly purified fraction was subjected to starch-gel electrophoresis for 16–18 h and then eluted from the gel, there was significant loss of heparin-cofactor activity but not of antithrombin activity. The electrophoretic patterns of the recovered proteins were not altered.

Effect of the carbon source and cyclic AMP on isocitrate dehydrogenase, succinate dehydrogenase, and malate dehydrogenase in Klebsiella pneumoniae C3

1982 ◽  
Vol 28 (10) ◽  
pp. 1101-1106 ◽  
Author(s):  
A. Juarez ◽  
R. Parés ◽  
J. Vives-Rego
Keyword(s):  
Klebsiella Pneumoniae ◽  
Stationary Phase ◽  
Succinate Dehydrogenase ◽  
Malate Dehydrogenase ◽  
Isocitrate Dehydrogenase ◽  
Specific Activity ◽  
Exponential Growth Phase ◽  
Specific Activities ◽  
In The Beginning ◽  
Excess Glucose

When strain C3 of Klebsiella pneumoniae is grown on a minimal medium with excess glucose, isocitrate dehydrogenase, malate dehydrogenase, and succinate dehydrogenase specific activities increase in the last period of the exponential growth phase and in the beginning of the stationary phase. Glucose exhaustion does not alter the development of malate dehydrogenase and succinate dehydrogenase, but specific activities are higher than those obtained with excess glucose. In contrast, glucose exhaustion can be correlated with a decrease of isocitrate dehydrogenase specific activity in the stationary phase. Induction of strain C3 isocitrate dehydrogenase by glucose in complex medium and repression by cAMP in mineral medium were observed. Glucose induction and the NADP/NADPH ratio are suggested as regulatory mechanisms controlling isocitrate dehydrogenase synthesis in the Enterobacteriaceae, but the former appears to be restricted to some Klebsiella strains.

IMPROVED PROCEDURES FOR PREPARATION AND CHARACTERIZATION OF MYROTHECIUM CELLULASE: PART 2. PURIFICATION PROCEDURES

1963 ◽  
Vol 41 (1) ◽  
pp. 671-696
Author(s):  
D. R. Whitaker ◽  
K. R. Hanson ◽  
P. K. Datta
Keyword(s):  
Ammonium Sulphate ◽  
Specific Activity ◽  
Deae Cellulose ◽  
Cellulase Activity ◽  
Starch Gel Electrophoresis ◽  
Polymethacrylic Acid ◽  
Citrate Buffer ◽  
Starch Gel ◽  
By Products

Two methods are described for purification of the cellulase of Myrothecium verrucaria from concentrated culture filtrates. The steps of method I are (1) fractionation with ammonium sulphate, (2) elution through Sephadex G25, (3) elution through Sephadex G75, (4) precipitation with polymethacrylic acid, and (5) elution through Amberlite CG50 with citrate buffer containing a gradient of urea concentration. The steps of method II are precipitation by saturated ammonium sulphate, (2) and (4) as above, elution through DEAE-cellulose with phosphate buffer containing 7 M urea, followed by (2) and (3) as above. The two methods gave final products with identical specific activities toward carboxymethyl cellulose; the increase in specific activity was approximately 12-fold. Starch-gel electrophoresis at pH 6.8 showed no major indications of heterogeneity. The purified enzyme was unstable but could be stored frozen in dilute salt solution.Enzyme passed through DEAE-cellulose without an inhibitor of cellulase activity was contaminated by DEAE-substituted oligoglucosides and subject to proteolysis. Urea could be replaced by cellobiose as an inhibitor but the latter gave enzyme contaminated by products of transfer reactions.Binding of urea by the resin is shown to influence significantly the resolution attainable in chromatographic fractionations on Amberlite CG-50 with buffers containing gradients of urea concentration.Procedures for dialysis and desalting of cellulases and a compact reaction vessel for pH stats are described.

IMPROVED PROCEDURES FOR PREPARATION AND CHARACTERIZATION OF MYROTHECIUM CELLULASE: PART 2. PURIFICATION PROCEDURES

1963 ◽  
Vol 41 (3) ◽  
pp. 671-696 ◽  
Author(s):  
D. R. Whitaker ◽  
K. R. Hanson ◽  
P. K. Datta
Keyword(s):  
Ammonium Sulphate ◽  
Specific Activity ◽  
Deae Cellulose ◽  
Cellulase Activity ◽  
Starch Gel Electrophoresis ◽  
Polymethacrylic Acid ◽  
Citrate Buffer ◽  
Starch Gel ◽  
By Products

Two methods are described for purification of the cellulase of Myrothecium verrucaria from concentrated culture filtrates. The steps of method I are (1) fractionation with ammonium sulphate, (2) elution through Sephadex G25, (3) elution through Sephadex G75, (4) precipitation with polymethacrylic acid, and (5) elution through Amberlite CG50 with citrate buffer containing a gradient of urea concentration. The steps of method II are precipitation by saturated ammonium sulphate, (2) and (4) as above, elution through DEAE-cellulose with phosphate buffer containing 7 M urea, followed by (2) and (3) as above. The two methods gave final products with identical specific activities toward carboxymethyl cellulose; the increase in specific activity was approximately 12-fold. Starch-gel electrophoresis at pH 6.8 showed no major indications of heterogeneity. The purified enzyme was unstable but could be stored frozen in dilute salt solution.Enzyme passed through DEAE-cellulose without an inhibitor of cellulase activity was contaminated by DEAE-substituted oligoglucosides and subject to proteolysis. Urea could be replaced by cellobiose as an inhibitor but the latter gave enzyme contaminated by products of transfer reactions.Binding of urea by the resin is shown to influence significantly the resolution attainable in chromatographic fractionations on Amberlite CG-50 with buffers containing gradients of urea concentration.Procedures for dialysis and desalting of cellulases and a compact reaction vessel for pH stats are described.

An International Collaborative Study to Investigate Standardisation of Hirudin Potency

1993 ◽  
Vol 69 (05) ◽  
pp. 430-435 ◽  
Author(s):  
Colin Longstaff ◽  
Man-Yu Wong ◽  
Patrick J Gaffney
Keyword(s):  
Specific Activity ◽  
Collaborative Study ◽  
Residual Activity ◽  
Quality Standard ◽  
Upper Limit ◽  
Assay Procedure ◽  
Specific Activities ◽  
International Collaborative ◽  
Range Of Values ◽  
International Collaborative Study

SummaryAn international collaborative study has been carried out to investigate the reproducibility of hirudin assays in 13 laboratories using four recombinant hirudins and one natural, sulphated product. A simple assay procedure was proposed involving the titration of α-thrombin with inhibitor and measurement of residual activity using a chromogenic substrate. A standard α-thrombin preparation was supplied to ensure that this reagent was of uniform quality throughout the study. The method appeared to present no difficulties and laboratories reported similar potencies for the 5 hirudin samples, in line with expected values. This gave 200–222 Thrombin Inhibitory Units/ampoule (TIU/ampoule) of lyophilised hirudin, with geometric coefficient of variation (gcv) values ranging from 10.15–15.97%. This corresponds to specific activities of approximately 14,300–15,900 TIU/mg protein. This is close to the upper limit of previously reported values of specific activity. We conclude that the precision of this determination compared with the wider range of values in the literature (8,000–16,000 thrombin inhibitory units [TIU]/mg) results from the use of good quality standard α-thrombin by all laboratories. This study has important implications for hirudin standardisation.

scholarly journals Gut Digestive Function and Microbiome after Correction of Experimental Dysbiosis in Rats by Indigenous Bifidobacteria

2021 ◽  
Vol 9 (3) ◽  
pp. 522
Author(s):  
Lyudmila V. Gromova ◽  
Elena I. Ermolenko ◽  
Anastasiya L. Sepp ◽  
Yulia V. Dmitrieva ◽  
Anna S. Alekseeva ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Digestive Enzymes ◽  
Specific Activity ◽  
Control Group ◽  
Enteropathogenic Escherichia Coli ◽  
Beneficial Bacteria ◽  
Opportunistic Bacteria ◽  
Dyspeptic Symptoms ◽  
Specific Activities ◽  
Impaired Motor Function

In recent years, great interest has arisen in the use of autoprobiotics (indigenous bacteria isolated from the organism and introduced into the same organism after growing). This study aimed to evaluate the effects of indigenous bifidobacteria on intestinal microbiota and digestive enzymes in a rat model of antibiotic-associated dysbiosis. Our results showed that indigenous bifidobacteria (the Bf group) accelerate the disappearance of dyspeptic symptoms in rats and prevent an increase in chyme mass in the upper intestine compared to the group without autoprobiotics (the C1 group), but significantly increase the mass of chyme in the colon compared to the C1 group and the control group (healthy animals). In the Bf group in the gut microbiota, the content of opportunistic bacteria (Proteus spp., enteropathogenic Escherichia coli) decreased, and the content of some beneficial bacteria (Bifidobacterium spp., Dorea spp., Blautia spp., the genus Ruminococcus, Prevotella, Oscillospira) changed compared to the control group. Unlike the C1 group, in the Bf group there was no decrease in the specific activities of maltase and alkaline phosphatase in the mucosa of the upper intestine, but the specific activity of maltase was decreased in the colon chyme compared to the control and C1 groups. In the Bf group, the specific activity of aminopeptidase N was reduced in the duodenum mucosa and the colon chyme compared to the control group. We concluded that indigenous bifidobacteria can protect the microbiota and intestinal digestive enzymes in the intestine from disorders caused by dysbiosis; however, there may be impaired motor function of the colon.

scholarly journals THE Ah PHENOTYPE. SURVEY OF FORTY-EIGHT RAT STRAINS AND TWENTY INBRED MOUSE STRAINS

Genetics ◽  
1982 ◽  
Vol 100 (1) ◽  
pp. 79-87
Author(s):  
Daniel W Nebert ◽  
Nancy M Jensen ◽  
Hisashi Shinozuka ◽  
Heinz W Kunz ◽  
Thomas J Gill
Keyword(s):  
Specific Activity ◽  
Inbred Mouse ◽  
Inbred Mouse Strain ◽  
Mouse Strains ◽  
Ah Receptor ◽  
Inbred Mouse Strains ◽  
Sprague Dawley ◽  
Rat Strains ◽  
Specific Activities ◽  
Inbred Rat

ABSTRACT Forty-four inbred and four randombred rat strains and 20 inbred mouse strains were examined for their Ah phenotype by determining the induction of liver microsomal aryl hydrocarbon (benzo[a]pyrene) hydroxylase activity (EC 1.14.14.1) by intraperitoneal treatment with either β-naphthoflavone or 3-methylcholanthrene. All 48 rat strains were found to be Ah-responsive. The maximally induced hydroxylase specific activities of the ALB/Pit, MNR/Pit, MR/Pit, SHR/Pit, and Sprague-Dawley strains were of the same order of magnitude as the basal hydroxylase specific activities of the ACI/Pit, F344/Pit, OKA/Pit, and MNR/N strains. Six of the 20 mouse strains were Ah-nonresponsive (i.e. lacking the normal induction response and presumably lacking detectable amounts of the Ah receptor). The basal hydroxylase specific activities of the BDL/N, NFS/N, STAR/N, and ST/JN mouse strains were more than twice as high as the maximally induced hydroxylase specific activity of the CBA/HT strain.——To date, 24 Ah-nonresponsive mouse strains have been identified, out of a total of 68 known to have been characterized. The reasons for not finding a single Ah-nonresponsive inbred rat strain—as compared with about one Ah-nonresponsive inbred mouse strain found for every three examined—remain unknown.

scholarly journals Regulation of lipogenic capacity in lactating rats

1982 ◽  
Vol 208 (3) ◽  
pp. 611-618 ◽  
Author(s):  
M R Grigor ◽  
A Geursen ◽  
M J Sneyd ◽  
S M Warren
Keyword(s):  
Mammary Gland ◽  
Fatty Acid ◽  
Malic Enzyme ◽  
Fatty Acid Synthase ◽  
Specific Activity ◽  
Mammary Glands ◽  
Lipogenic Enzymes ◽  
Pregnancy And Lactation ◽  
Specific Activities

1. The rate of mammary-gland lipogenesis measured in vivo from 3H2O was suppressed after decreasing the milk demand by decreasing the number of pups from ten to two or three, as well as by giving diets containing lipid [Grigor & Warren (1980) Biochem. J. 188, 61-65]. 2. The specific activities of the lipogenic enzymes fatty acid synthase, glucose 6-phosphate dehydrogenase and ‘malic’ enzyme increased between 6- and 10-fold in the mammary gland and between 2- and 3-fold in the livers during the first 10 days of lactation. The increases in specific activity coupled with the doubling of liver mass which occurred during pregnancy and lactation resulted in considerable differences in total liver activities when compared with virgin animals. 3. Although consumption of a diet containing 20% peanut oil suppressed the activities of the three lipogenic enzymes in the livers, only the ‘malic’ enzyme was affected in the mammary glands. 4. In contrast, decreased milk demand did not affect the specific activities of any of the liver enzymes, whereas it resulted in suppression of all three lipogenic enzymes of the mammary glands. There was no effect on either the cytoplasmic malate dehydrogenase or the lactate dehydrogenase of the mammary gland. 5. In all the experiments performed, the activity of the fatty acid synthase correlated with the amount of material precipitated by the rabbit antibody raised against rat fatty acid synthase.

scholarly journals A chromatographic preparation of ox growth hormone

1966 ◽  
Vol 100 (3) ◽  
pp. 593-600 ◽  
Author(s):  
M Wallis ◽  
HBF Dixon
Keyword(s):  
Molecular Weight ◽  
Growth Hormone ◽  
Gel Electrophoresis ◽  
Anterior Pituitary ◽  
Gel Filtration ◽  
Deae Cellulose ◽  
Starch Gel Electrophoresis ◽  
Highly Active ◽  
Cross Linkage ◽  
Starch Gel

1. A method is described for the chromatographic preparation of ox growth hormone. It involves chromatography of an extract of anterior pituitary lobes on DEAE-cellulose, followed by rechromatography on a dextran gel of low cross-linkage (Sephadex G-100). 2. The product is highly active in growth-hormone assays, and is obtained in good yield. It was homogeneous by several criteria, but showed some heterogeneity on starch-gel electrophoresis. 3. The molecular weight of the hormone was estimated from its behaviour on gel-filtration columns under various conditions. Evidence that the hormone may dissociate into sub-units under some conditions is presented.

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