scholarly journals Studies on bone enzymes. Distribution of acid hydrolases, alkaline phenylphosphatase, cytochrome oxidase and catalase in subcellular fraction of bone tissue homogenates

1965 ◽  
Vol 97 (2) ◽  
pp. 389-392 ◽  
Author(s):  
G Vaes ◽  
P Jacques
Keyword(s):  
Cytochrome Oxidase ◽  
Subcellular Fraction ◽  
Microsomal Fraction ◽  
Specific Activity ◽  
Distribution Patterns ◽  
Mitochondrial Fraction ◽  
Acid Hydrolases ◽  
Mitochondrial Fractions ◽  
Tissue Homogenates ◽  
Isopycnic Centrifugation

1. When bone homogenates were fractionated according to the scheme developed for liver by de Duve, Pressman, Gianetto, Wattiaux & Appelmans (1955), all the enzymes assayed except cytochrome oxidase were found to occur partly in soluble and partly in particulate fractions. Among the particle-bound enzymes, the highest specific activity was found in the heavy-mitochondrial fraction for cytochrome oxidase, in the microsomal fraction for alkaline phenylphosphatase and in the light-mitochondrial fraction for eight acid hydrolases and for catalase. 2. Combined heavy-mitochondrial and light-mitochondrial fractions were subfractionated by isopycnic centrifugation in density gradients of sucrose or glycogen. In the various systems tried, cytochrome oxidase showed a relatively narrow distribution range with a sharp peak; the acid hydrolases and catalase showed flat and irregular distribution patterns, differing slightly in shape from one enzyme to the other. However, it was not possible to achieve a marked separation between the various enzymes under study. 3. It is concluded from these results that the acid hydrolases belong to special cytoplasmic particles, probably lysosomes, and that these particles are physically and enzymically heterogeneous. Catalase appears to be non-mitochondrial and could also belong to the lysosomes; but the possibility of an association with another type of particle must be kept in mind in view of what is known of liver catalase. Alkaline phenylphosphatase is largely attached to microsomal elements.

Mitochondrial number, cytochrome oxidase and succinic dehydrogenase activity in Xenopus laevis oocytes

Development ◽  
1981 ◽  
Vol 62 (1) ◽  
pp. 395-409
Author(s):  
E. Marinos ◽  
F. S. Billett
Keyword(s):  
Xenopus Laevis ◽  
Cytochrome Oxidase ◽  
Specific Activity ◽  
Succinic Dehydrogenase ◽  
Mitochondrial Fraction ◽  
Xenopus Laevis Oocytes ◽  
Mitochondrial Fractions ◽  
Rate Of Increase ◽  
Mitochondrial Cloud ◽  
Mitochondrial Number

An estimate has been made of the numbers of mitochondria in the mitochondrial cloud (Balbiani body) of Xenopus laevis oocytes ranging in size from 50 to 250 μm. The mitochondrial number is expressed in terms of a ‘standard ’ organelle measuring 2 μm in length and 0·2 μm in diameter and is derived by measurements on electron micrographs of sections through the cloud. It is found that the amount of mitochondrial material rises very rapidly as the oocyte grows in size. At the time the cloud disperses, in oocytes of about 300 μm in diameter, it is estimated that there are the equivalent of over 500000 mitochondria in each cell. The rate of increase is very similar to the rate of accumulation of mitochondrial DNA during the same period of growth. Using a polarographic technique the specific activity of cytochrome oxidase and succinic dehydrogenase was determined in mitochondrial fractions isolated from oocytes over a size range of 80–1200 μm in diameter. Although the specific activity of succinic dehydrogenase remains constant that of cytochrome oxidase falls sharply during the period when the mitochondria are replicating rapidly, i.e. up to about 300 μm diameter. In larger oocytes the specific activity of enzymes appears to remain constant but increasing contamination of the isolated mitochondrial fraction does not allow conclusions to be drawn from the enzyme loading of the mitochondria once they have dispersed from the cloud. The results are discussed in relation to the possibility that mitochondrial replication preceeds, or at least outpaces, mitochondrial differentiation during the course of oogenesis.

Subcellular distribution of renal tripeptide-releasing exopeptidases active on collagen-like sequences

1987 ◽  
Vol 252 (5) ◽  
pp. F890-F898
Author(s):  
K. J. Andersen ◽  
J. K. McDonald
Keyword(s):  
Subcellular Distribution ◽  
Microsomal Fraction ◽  
Specific Activity ◽  
Rat Kidney ◽  
Broad Band ◽  
Distribution Patterns ◽  
Kidney Cortex ◽  
Rat Kidney Cortex ◽  
Tripeptidyl Peptidase ◽  
N Terminus

The rat kidney cortex was found to contain two N-terminal exopeptidases of the tripeptidyl peptidase (TPP) class. Each required a free N-terminus to catalyze the release of collagen-related (Gly-Pro-X) "triplets." In accordance with their apparent pH optima, activities were routinely determined fluorimetrically at pH 4.0 (TPP 4) and at pH 7.0 (TPP 7) on Gly-Pro-Met-2-naphthylamide. The specific activity in both the homogenate and the classical subfractions was much greater at pH 7 than at pH 4. Subfractionation of the microsomal fraction by equilibrium banding in sucrose did not separate the TPP 4 and TPP 7 activities. The banding density (1.18 g/ml) and the distribution patterns for TPP 7 in the microsomal subfractions, and also in the subfractions of the small lysosomes in the mitochondrial-lysosomal (ML) fraction, demonstrate that TPP 7 is associated with smooth membranes. The TPP 4 and TPP 7 activities were clearly separated during subfractionation of the ML fraction. Rate sedimentation demonstrated that TPP 4 was present in the large, fast-sedimenting lysosomes (protein droplets) and in a heterogeneous broad band of smaller lysosomes. Equilibrium banding of the small lysosomes gave two distinct TPP 4-containing populations at densities 1.20 and 1.235 g/ml. Notably, dipeptidyl peptidase II (DPP II) gave identical banding densities and showed distributions very similar to TPP 4.(ABSTRACT TRUNCATED AT 250 WORDS)

Phosphoribosyl Pyrophosphate Degradation in Human Tissues

1974 ◽  
Vol 52 (12) ◽  
pp. 1162-1166 ◽  
Author(s):  
Irving H. Fox ◽  
Pamela J. Marchant
Keyword(s):  
Microsomal Fraction ◽  
Specific Activity ◽  
Maximum Activity ◽  
Human Tissues ◽  
Phosphoribosyl Pyrophosphate ◽  
Phosphorylated Compounds ◽  
Tissue Homogenates ◽  
Sugar Derivatives ◽  
Hydrolysis Of ◽  
Alkaline Range

Enzymatic hydrolysis of phosphoribosyl pyrophosphate (PP-ribose-P) in dialyzed human tissue homogenates had a specific activity ranging from 0.15 to 1.37 μmol mg−1 h−1. Our observations on human placenta have characterized this reaction: (1) PP-ribose-P was degraded in the absence of Mg with stoichiometric release of Pi at 37 °C; (2) the reaction occurred from pH 5 to 10.5 with the maximum activity in the alkaline range; (3) PP-ribose-P degrading activity was localized mainly in the microsomal fraction; (4) alkaline phosphatase rather than 5′-nucleotidase was responsible for the degradation of PP-ribose-P; (5) the Km for PP-ribose-P was 0.3 mM; (6) PP-ribose-P hydrolysis was altered by many phosphorylated compounds, both nucleoside and sugar derivatives, Pi, and PPi in concentrations ranging from 0.1 to 10.0 mM.

DISTRIBUTION OF CHOLINESTERASE IN NORMAL HUMAN MUSCLE

1963 ◽  
Vol 41 (8) ◽  
pp. 1713-1720 ◽  
Author(s):  
J. Crispin Smith ◽  
Vera M. Foldes ◽  
Francis F. Foldes
Keyword(s):  
Skeletal Muscle ◽  
Microsomal Fraction ◽  
Cholinesterase Activity ◽  
Specific Activity ◽  
Mitochondrial Fraction ◽  
Human Muscle ◽  
Ventricular Muscle ◽  
Cardiac Ventricular Muscle ◽  
Normal Human ◽  
Quadriceps Femoris Muscles

The intracellular distribution of acetylcholinesterase and butyrylcholinesterase in biopsied human rectus abdominis and quadriceps femoris muscles was investigated. Differential ultracentrifugal fractionation showed that the highest specific activity of acetylcholinesterase was associated with the microsomal fraction, this enzyme being concentrated 19-fold compared to the original homogenate. The highest specific activity of butyrylcholinesterase was found in the non-particulate fraction. The mitochondrial fraction showed the lowest specific cholinesterase activity. Wide variation of cholinesterase activity was found in various autopsied muscles, the highest being oculomotor muscle and the lowest cardiac ventricular muscle. About 90% of the cholinesterase activity of skeletal muscle was due to acetylcholinesterase and 10% to butyrylcholinesterase.

Seasonal and starvation-induced changes in enzymatic pattern of frog nephron

1961 ◽  
Vol 201 (5) ◽  
pp. 781-785 ◽  
Author(s):  
Morris J. Karnovsky ◽  
S. Ralph Himmelhoch
Keyword(s):  
Cytochrome Oxidase ◽  
Enzyme Activities ◽  
Specific Activity ◽  
Hydrolytic Enzyme ◽  
Succinic Dehydrogenase ◽  
Krebs Cycle ◽  
Hexose Monophosphate ◽  
Hexose Monophosphate Shunt ◽  
Mitochondrial Fractions ◽  
Proximal Tubular

Oxidative and hydrolytic enzyme activities in the nephron of the frog were investigated using histochemical and spectrophotometric techniques. The distal tubule was found to possess high activities of enzymes of glycolysis and the Krebs cycle, as well as high cytochrome oxidase activity, but had only slight activities of the hexose-monophosphate shunt enzymes. The proximal tubule had high activities of glycolytic enzymes, enzymes of the hexose-monophosphate shunt, and enzymes of the Krebs cycle, except for succinic dehydrogenase, which was present in very low activity. Proximal tubular cytochrome oxidase was found to vary according both to the season and to environ mental conditions. In freshly caught summer frogs it was present in high activity, but in winter frogs or summer frogs starved for periods of 10–30 days, no proximal tubular cytochrome oxidase could be detected, although other proximal tubular enzyme activities remained unchanged as far as could be histochemically determined. The specific activity of cytochrome oxidase in mitochondrial fractions was found to decrease by about 50% in the kidneys of starved and winter frogs, as compared with summer frogs.

scholarly journals Biogenesis of mitochondria. Phospholipid synthesis in vitro by yeast mitochondrial and microsomal fractions

1974 ◽  
Vol 144 (2) ◽  
pp. 265-275 ◽  
Author(s):  
G S Cobon ◽  
P D Crowfoot ◽  
A W Linnane
Keyword(s):  
Phosphatidic Acid ◽  
Microsomal Fraction ◽  
De Novo ◽  
Specific Activity ◽  
Neutral Lipids ◽  
Mitochondrial Fraction ◽  
Biogenesis Of Mitochondria ◽  
Glycerolipid Synthesis ◽  
Phosphatidylcholine Synthesis

The ability in vitro of yeast mitochondrial and microsomal fractions to synthesize lipid de novo was measured. The major phospholipids synthesized from sn-[2-3H]glycerol 3-phosphate by the two microsomal fractions were phosphatidylserine, phosphatidylinositol and phosphatidic acid. The mitochondrial fraction, which had a higher specific activity for total glycerolipid synthesis, synthesized phosphatidylglycerol, phosphatidylethanolamine, phosphatidylinositol, phosphatidylserine and phosphatidic acid, together with smaller amounts of neutral lipids and diphosphatidylglycerol. Phosphatidylcholine synthesis from both S-adenosyl[Me-14C]methionine and CDP-[Me-14C]choline appeared to be localized in the microsomal fraction.

Biosynthesis of Phosphatidic Acid in Mitochondria and Microsomes Via the Acylation of sn-Glycero-3-phosphate

1972 ◽  
Vol 50 (8) ◽  
pp. 936-948 ◽  
Author(s):  
J. B. Davidson ◽  
N. Z. Stanacev
Keyword(s):  
Phosphatidic Acid ◽  
Subcellular Fraction ◽  
Microsomal Fraction ◽  
Specific Activity ◽  
Marker Enzymes ◽  
Acyltransferase Activity ◽  
Isolated Mitochondria ◽  
Direct Acylation ◽  
Submitochondrial Fractions ◽  
Nadph Cytochrome C Reductase

Nuclei-free homogenate, prepared from guinea pig livers, was fractionated into subcellular particles which were then examined for the activities of two microsomal marker enzymes, glucose-6-phosphatase and NADPH: cytochrome c reductase. In an incubation system containing sn-glycero-3-phosphate, fatty acid, and various cofactors the intracellular distribution of acyl-CoA: sn-glycero-3-phosphate acyltransferase(s) was studied and compared with the distribution of the two microsomal marker enzymes.Results obtained showed that the highest specific activity for the acylation of sn-glycero-3-phosphate was associated with the microsomal fraction and the activity in each subcellular fraction paralleled activities of the two microsomal marker enzymes. Furthermore, the amount of acyl-CoA: sn-glycero-3-phosphate acyltransferase activity observed in the mitochondrial and submitochondrial fractions could be accounted for by the content of endoplasmic reticulum as determined by the marker enzymes. This observation was also true for brain, heart, and kidney, as well as for rat liver.These results are interpreted as evidence that isolated mitochondria are unable to synthesize phosphatidic acid by direct acylation of sn-glycero-3-phosphate.

scholarly journals Cytochrome c oxidase is preferentially synthesized in the rough endoplasmic reticulum-mitochondrion complex in rat liver

1982 ◽  
Vol 208 (2) ◽  
pp. 505-507 ◽  
Author(s):  
S Parimoo ◽  
N Rao ◽  
G Padmanaban
Keyword(s):  
Endoplasmic Reticulum ◽  
Cytochrome C ◽  
Cytochrome Oxidase ◽  
Cytochrome C Oxidase ◽  
Rat Liver ◽  
Rough Endoplasmic Reticulum ◽  
Specific Activity ◽  
Mitochondrial Fraction

The specific activity and content of cytochrome oxidase in the rough endoplasmic reticulum-mitochondrion complex are higher than in the mitochondrial fraction. Radiolabelling studies with the use of hepatocytes and isolated microsomal and rough endoplasmic reticulum-mitochondrion fractions, followed by immunoprecipitation with anti-(cytochrome oxidase) antibody, reveal that the nuclear-coded cytoplasmic subunits of cytochrome oxidase are preferentially synthesized in the latter fraction. The results have a bearing on the mechanism of transport of these subunits into mitochondria.

scholarly journals Enzymatically inactive forms of acetyl-CoA carboxylase in rat liver mitochondria

1988 ◽  
Vol 251 (3) ◽  
pp. 881-885 ◽  
Author(s):  
J B Allred ◽  
C R Roman-Lopez
Keyword(s):  
Rat Liver ◽  
Microsomal Fraction ◽  
Specific Activity ◽  
Polyacrylamide Gel Electrophoresis ◽  
Mitochondrial Fraction ◽  
Liver Mitochondria ◽  
Rat Liver Mitochondria ◽  
Outer Mitochondrial Membrane ◽  
Acetyl Coa Carboxylase ◽  
Acetyl Coa

Biotinyl proteins were labelled by incubation of SDS-denatured preparations of subcellular fractions of rat liver with [14C]methylavidin before polyacrylamide-gel electrophoresis. Fluorographic analysis showed that mitochondria contained two forms of acetyl-CoA carboxylase [acetyl-CoA:carbon dioxide ligase (ADP-forming) EC 6.4.1.2], both of which were precipitated by antibody to the enzyme. When both forms were considered, almost three-quarters of the total liver acetyl-CoA carboxylase was found in the mitochondrial fraction of liver from fed rats while only 3.5% was associated with the microsomal fraction. The remainder was present in cytosol, either as the intact active enzyme or as a degradation product. The actual specific activity of the cytosolic enzyme was approx. 2 units/mg of acetyl-CoA carboxylase protein while that of the mitochondrial enzyme was about 20-fold lower, indicating that mitochondrial acetyl-CoA carboxylase was relatively inactive. Fractionation of mitochondria with digitonin showed that acetyl-CoA carboxylase was associated with the outer mitochondrial membrane. The available evidence suggests that mitochondrial acetyl-CoA carboxylase represents a reservoir of enzyme which can be released and activated under lipogenic conditions.

DISTRIBUTION OF CHOLINESTERASE IN NORMAL HUMAN MUSCLE

1963 ◽  
Vol 41 (1) ◽  
pp. 1713-1720 ◽  
Author(s):  
J. Crispin Smith ◽  
Vera M. Foldes ◽  
Francis F. Foldes
Keyword(s):  
Skeletal Muscle ◽  
Microsomal Fraction ◽  
Cholinesterase Activity ◽  
Specific Activity ◽  
Mitochondrial Fraction ◽  
Human Muscle ◽  
Ventricular Muscle ◽  
Cardiac Ventricular Muscle ◽  
Normal Human ◽  
Quadriceps Femoris Muscles

The intracellular distribution of acetylcholinesterase and butyrylcholinesterase in biopsied human rectus abdominis and quadriceps femoris muscles was investigated. Differential ultracentrifugal fractionation showed that the highest specific activity of acetylcholinesterase was associated with the microsomal fraction, this enzyme being concentrated 19-fold compared to the original homogenate. The highest specific activity of butyrylcholinesterase was found in the non-particulate fraction. The mitochondrial fraction showed the lowest specific cholinesterase activity. Wide variation of cholinesterase activity was found in various autopsied muscles, the highest being oculomotor muscle and the lowest cardiac ventricular muscle. About 90% of the cholinesterase activity of skeletal muscle was due to acetylcholinesterase and 10% to butyrylcholinesterase.

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