Zymographic approach to determine the intrinsic enzyme specific activity of diamine oxidase in presence of interfering enzymes

2017 ◽  
Vol 975 ◽  
pp. 78-85 ◽  
Author(s):  
Samaneh Ahmadifar ◽  
Tien Canh Le ◽  
Lucia Marcocci ◽  
Paola Pietrangeli ◽  
Mircea Alexandru Mateescu
Keyword(s):  
Diamine Oxidase ◽  
Specific Activity ◽  
Enzyme Specific Activity

Comparative studies of activity and properties of ferredoxin–NADP+ reductase during cold hardening of wheat

1976 ◽  
Vol 54 (16) ◽  
pp. 1896-1902 ◽  
Author(s):  
Joseph Riov ◽  
Gregory N. Brown
Keyword(s):  
Activation Energy ◽  
Spring Wheat ◽  
Comparative Studies ◽  
Low Temperatures ◽  
Specific Activity ◽  
Cold Hardening ◽  
Heat Inactivation ◽  
Enzyme Molecule ◽  
Enzyme Specific Activity ◽  
Michaelis Constants

Activity and properties of chloroplast ferredoxin–NADP− reductase (EC 1.6.7.1) were studied during cold hardening of two varieties of wheat (Triticum aestivnm), hardy Kharkov (winter wheat) and much less hardy Rescue (spring wheat), to determine whether adaptation to low temperatures involves changes in the activity and properties of this enzyme. Specific activity of ferredoxin–NADP− reductase increased during hardening of both varieties, but the increase was much greater in the more hardy variety, Kharkov 22 MC. No changes were found in the Michaelis constants for NADPH and 2,6-dichlorophenol indophenol, activation energy values, inhibition constants for p-chloromercuriphenylsulfonate, and sensitivity toward cold and heat inactivation of the enzyme from control and cold-hardened seedlings of both varieties. The data suggest that there is a preferential synthesis of ferredoxin–NADP− reductase during hardening of wheat, but the enzyme molecule remains unchanged.

scholarly journals Mitochondrial development in liver of foetal and newborn rats

1971 ◽  
Vol 121 (2) ◽  
pp. 341-347 ◽  
Author(s):  
S. Jakovcic ◽  
J. Haddock ◽  
G. S. Getz ◽  
M. Rabinowitz ◽  
H. Swift
Keyword(s):  
Specific Activity ◽  
Liver Mitochondria ◽  
Postnatal Period ◽  
Neonatal Rat ◽  
Inner Mitochondrial Membrane ◽  
Adult Liver ◽  
Respiratory Enzymes ◽  
Respiratory Enzyme ◽  
Foetal Liver ◽  
Enzyme Specific Activity

The development of the inner mitochondrial membrane in foetal and neonatal rat liver was studied by following three parameters: (1) the activity of several respiratory enzymes in homogenates and purified mitochondria, (2) the spectrophotometric determination of cytochrome content in the mitochondria and (3) the cardiolipin content in both homogenates and purified mitochondria. Respiratory-enzyme activities of homogenates of foetal liver were one-quarter to one-twentieth of those of homogenates of adult liver, and the enzyme specific activities in purified mitochondria from foetal liver were one-half to one-eighth of those in mitochondria from adult liver. The cardiolipin content of liver homogenates increased approximately twofold during the development period, but there was no significant change in the cardiolipin content of purified mitochondria. It is concluded that cell mitochondrial content approximately doubles in the immediate postnatal period. There was no evidence for an increase in the relative amount of cristae protein in mitochondria during this period to account for increases in mitochondrial enzyme specific activity, since cardiolipin and cytochrome concentrations remained unchanged and electron micrographs revealed no differences. The cause of the lower respiratory-enzyme specific activity in foetal liver mitochondria is unclear. Qualitative differences in respiratory units in foetal and mature animals are suggested.

scholarly journals Regulation of fitness in yeast overexpressing glycolytic enzymes: parameters of growth and viability

1992 ◽  
Vol 59 (1) ◽  
pp. 35-48 ◽  
Author(s):  
R. F. Rosenzweig
Keyword(s):  
Growth Rate ◽  
Stationary Phase ◽  
Specific Activity ◽  
Current Model ◽  
Carbon Sources ◽  
Fold Increase ◽  
Culture Cell ◽  
Phase Density ◽  
Enzyme Specific Activity ◽  
High Copy Plasmid

SummaryCurrent models predict that large increases over wild-type in the activity of one enzyme will not alter an organism's fitness. This prediction is tested in Saccharomyces cerevisiae through the use of a high copy plasmid that bears one of the following: hexokinase B (HEXB), phosphoglucose isomerase (PGI), phosphofructokinase (PFKAandPFKB), or pyruvate kinase (PYK). Transformants containing these plasmids demonstrate a four to ten-fold increase in enzyme specific activity over either the parent strain or transformants containing the plasmid alone. Haploid and diploid transformants derived from independent backgrounds were grown on both fermentable and non-fermentable carbon sources and evaluated for several components of fitness. These include growth rate under non-limiting conditions, maximum stationary phase density, and viability in extended batch culture. Cell viability is not affected by overproduction of these enzymes. Growth rate and stationary phase density do not differ significantly among strains that overexpressHEXB, PGIor contain the vector alone.PFKA, Btransformants show reduced growth rate on glucose in one background only. For these loci the current model is confirmed. By contrast, when grown on glucose, yeast overexpressingPYKdemonstrate reduced growth rate and increased stationary phase density in both backgrounds. These effects are abolished in cells containing plasmids with a Tn5 disrupted copy of thePYKgene. Our results are consistent with reports that the PYK locus may exert control over the yeast cell cycle and suggest that it will be challenging to model relations between fitness and activity for multifunctional proteins.

scholarly journals Optimum conditions for isoamylase production by Pseudomonas sp.

2013 ◽  
Vol 7 (1) ◽  
pp. 3-10
Author(s):  
Hameed M. Jasim ◽  
Haneen S. Abdul-Wahab
Keyword(s):  
Batch Culture ◽  
Specific Activity ◽  
Cultural Factors ◽  
Pseudomonas Sp ◽  
Production Medium ◽  
Optimum Conditions ◽  
Medium Ph ◽  
Enzyme Specific Activity ◽  
Initial Medium

Different nutritional and cultural factors were studied to determine the optimum conditions for isoamylase production by Pseudomonas sp. in a batch culture of the production medium. These factors include carbon, nitrogen and phosphate sources and their concentrations, temperature and pH. Results showed that the optimum conditions for isoamylase production by Pseudomonas sp. were achieved when the production medium was supplemented with maltose 1%, peptone 0.4%, and K2HPO4 0.4% as a carbon, nitrogen, and phosphate sources respectively, at initial medium pH 6, and incubation at 28°C for 24 hours. Under these conditions isoamylase productivity reaches the maximum, at which enzyme specific activity was 0.85 U/mg proteins.

scholarly journals DIFFERENTIATION OF ENDOPLASMIC RETICULUM IN HEPATOCYTES

1971 ◽  
Vol 49 (2) ◽  
pp. 264-287 ◽  
Author(s):  
A. Leskes ◽  
P. Siekevitz ◽  
G. E. Palade
Keyword(s):  
Endoplasmic Reticulum ◽  
Phosphatase Activity ◽  
Rough Endoplasmic Reticulum ◽  
Specific Activity ◽  
Smooth Endoplasmic Reticulum ◽  
Rat Hepatocytes ◽  
Adult Level ◽  
Enzyme Specific Activity ◽  
Cytochemical Reaction ◽  
And Control

The distribution of glucose-6-phosphatase activity in rat hepatocytes during a period of rapid endoplasmic reticulum differentiation (4 days before birth-1 day after birth) was studied by electron microscope cytochemistry. Techniques were devised to insure adequate morphological preservation, retain glucose-6-phosphatase activity, and control some other possible artifacts. At all stages examined the lead phosphate deposited by the cytochemical reaction is localized to the endoplasmic reticulum and the nuclear envelope. At 4 days before birth, when the enzyme specific activity is only a few per cent of the adult level, the lead deposit is present in only a few hepatocytes. In these cells a light deposit is seen throughout the entire rough-surfaced endoplasmic reticulum. At birth, when the specific activity of glucose-6-phosphatase is approximately equal to that of the adult, nearly all cells show a positive reaction for the enzyme and, again, the deposit is evenly distributed throughout the entire endoplasmic reticulum. By 24 hr postparturition all of the rough endoplasmic reticulum, and in addition the newly formed smooth endoplasmic reticulum, contains heavy lead deposits; enzyme activity at this stage is 250% of the adult level. These findings indicate that glucose-6-phosphatase develops simultaneously within all of the rough endoplasmic reticulum membranes of a given cell, although asynchronously in the hepatocyte population as a whole. In addition, the enzyme appears throughout the entire smooth endoplasmic reticulum as the membranes form during the first 24 hr after birth. The results suggest a lack of differentiation within the endoplasmic reticulum with respect to the distribution of glucose-6-phosphatase at the present level of resolution.

A quantitative study of the fixation of acid phosphatase by formaldehyde and its relevance to histochemistry

1974 ◽  
Vol 186 (1083) ◽  
pp. 137-164 ◽  
Keyword(s):  
Acid Phosphatase ◽  
Specific Activity ◽  
Relative Activity ◽  
Buffer System ◽  
Inhibitory Effects ◽  
Fixed Tissue ◽  
Apparent Loss ◽  
Nitrophenyl Phosphate ◽  
Enzyme Specific Activity ◽  
Tissue Nitrogen

Fixatives reduce the activities of most enzymes in sections and blocks of tissue, but pre­vious studies do not agree by how much. In this paper it is shown that the disagreement is due principally to the use of inappropriate measurement parameters and to variations in the way tissues are prepared. Some new approaches to this problem are reported. They are illustrated by measurements on the activities of acid phosphatase (against p -nitrophenyl phosphate) remaining in blocks and cryostat-cut sections of hamster kidney after fixation in buffered solutions of formaldehyde. Whole fresh kidneys were cut into 1 mm 3 cubes. Two or more ‘lots’ of 9-12 cubes were chosen at random, fixed, washed, homogenized and assayed for residual enzyme specific activity which was expressed in terms of tissue nitrogen content or frozen-dried mass rather than protein content. The activity apparently remaining after fixation depends on, it was found, the time and temperature of fixation, the buffer salt in the fixative, the length of the post-fixation wash, and the duration and temperature of homogenization. For example, the relative activity (that is, specific activity expressed as a percentage of that of unfixed tissue) of acid phos­phatase in cubes assayed immediately after fixation is 46%; after a 24 h post-fixation wash in cold buffer it rises to 84%. Fixation causes only a 2% loss of nitrogenous material from the cubes. A further 2.7% is lost during the 24 h wash. Thus, much of the apparent loss of enzyme specific activity in fixed tissue is due to the inhibitory effects of fixative remaining in the tissue. Similar inhibition effects were observed in cryostat-cut sections, although sections frozen-dried on to coverslips showed a greater tolerance than fresh ones. For example, after 5 min fixation and without a post-fixation wash, the activity of acid phosphatase in fresh, cryostat-cut, mounted sections falls by about 30%, but that in frozen-dried ones is un­affected. After 25 min fixation, however the activity in the frozen-dried sections drops to about 84% of that of the unfixed tissue. A nitrogen-free buffer system, 5-norbornene-2, 3-dicarboxylic acid-NaOH, was used as the fixative vehicle for much of this study. A higher acid phosphatase activity is retained in tissues fixed or stored in this buffer than in either cacodylate or phosphate.

Purified postheparin plasma lipoprotein lipase in primary hyperlipoproteinemias

1975 ◽  
Vol 39 (6) ◽  
pp. 1022-1033 ◽  
Author(s):  
D. Ganesan ◽  
R. H. Bradford ◽  
G. Ganesan ◽  
W. J. McConathy ◽  
P. Alaupovic ◽  
...  
Keyword(s):  
Lipoprotein Lipase ◽  
Specific Activity ◽  
Lipolytic Activity ◽  
Plasma Lipoprotein ◽  
Type I ◽  
Activity Type ◽  
Type Iii ◽  
Protein Peak ◽  
Enzyme Specific Activity ◽  
Hydrolysis Of

Purified postheparin plasma lipoprotein lipase (LPL) of normolipidemic and primary hyperlipoproteinemic subjects was characterized by lipoprotein C polypeptide activation and specificity for triglycerides in chylomicrons and VLDL. Chromatography of normal LPL on Sephadex G-100 resulted in two protein peaks, LPLC-1 (activated by C-I but not C-II) and LPLC-II (activated by C-II but not C-I). LPL from type I hyperlipoproteinemic subjects was not activated by C-I and C-II activation was reduced to 40% of control. Hydrolysis of chylomicron and VLDL triglycerides was severely impaired. Although chromatography of type I LPL resulted in two protein peaks, the protein peak corresponding to LPLC-I did not exhibit lipolytic activity and LPLC-II was reduced to 50% of control in protein and enzyme specific activity. Type III LPL was normal in respect to LPLC-I while LPLC-II averaged 40% of control. Hydrolysis of chylomicron and VLDL was reduced to 50% and 10% of control, respectively. An etiological implication for LPLC-I and/or LPLC-II in type I and III hyperlipoproteinemias is suggested.

Distribution of the multiple molecular forms of glucose-6-phosphate dehydrogenase in different physiological states

1979 ◽  
Vol 57 (5) ◽  
pp. 396-401 ◽  
Author(s):  
Hsiao-Lin Chang ◽  
Darold Holten ◽  
Rom Karin
Keyword(s):  
Enzyme Activity ◽  
Mammary Gland ◽  
Rat Liver ◽  
Specific Activity ◽  
Molecular Forms ◽  
Polyacrylamide Gels ◽  
Multiple Molecular Forms ◽  
Pregnancy And Lactation ◽  
Enzyme Specific Activity ◽  
Glucose 6 Phosphate Dehydrogenase

The distribution of the multiple molecular forms of rat liver and mammary gland glucose-6-phosphate dehydrogenase was determined by electrophoresis on 5% polyacrylamide gels. In both of these organs, changes in the distribution of enzyme activity among the several forms was slight even when approximately 20- to 40-fold changes in enzyme specific activity were achieved by fasting-refeeding experiments (for liver) or during pregnancy and lactation (for mammary gland), it was concluded that the induction of glucose-6-phosphate dehydrogenase in these two organs occurs without any major redistribution among the multiple molecular forms of this enzyme.

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