scholarly journals Hepatic Golgi fractions resolved into membrane and content subfractions.

1982 ◽  
Vol 92 (3) ◽  
pp. 822-832 ◽  
Author(s):  
K E Howell ◽  
G E Palade
Keyword(s):  
Electron Microscopy ◽  
Rat Liver ◽  
Enzyme Activities ◽  
Degree Of Contamination ◽  
Golgi Membranes ◽  
Membrane Enzyme ◽  
Biochemical Assays ◽  
The Reformation ◽  
Liver Homogenates

Golgi fractions isolated from rat liver homogenates have been resolved into membrane and content subfractions by treatment with 100 mM Na2CO3 pH 11.3. This procedure permitted extensive extraction of content proteins and lipoproteins, presumably because it caused an alteration of Golgi membranes that minimized the reformation of closed vesicles. The type and degree of contamination of the fractions was assessed by electron microscopy and biochemical assays. The membrane subfraction retained 15% of content proteins and lipids, and these could not be removed by various washing procedures. The content subfraction was contaminated by both membrane fragments and vesicles and accounted for 5 to 10% of the membrane enzyme activities of the original Golgi fraction. The lipid compositions of the subfractions was determined, and the phospholipids of both membrane and content were found to be uniformly labeled with [33P]phosphate administered in vivo.

scholarly journals Presence of nadph-cytochrome P-450 reductase in rat liver golgi membranes: Evidence obtained by immunoadsorption method

1978 ◽  
Vol 79 (2) ◽  
pp. 590-597 ◽  
Author(s):  
A Ito ◽  
GE Palade
Keyword(s):  
Rat Liver ◽  
Reductase Activity ◽  
Low Density Lipoprotein ◽  
Density Lipoprotein ◽  
Nadph Cytochrome ◽  
Golgi Membranes ◽  
Cyt C ◽  
Bona Fide ◽  
Liver Homogenates ◽  
Cytochrome P 450

Light Golgi fractions (GF(1+2)) prepared from rat liver homogenates by a modification of the Ehrenreich et al. procedure (J. Cell Biol. 59:45) had significant NADPH-cytochrome P(450) reductase (NADPH-cyt c reductase) activity if assayed immediately after their isolation. An antibody raised in rabbits against purified microsomal and Golgi fractions. To find out whether this activity is located in bona fide Golgi elements or in contaminating microsomal vesicles, we used the following 3-step immunoadsorption procedure: (a) antirabbit IgG (raised in goats) was conjugated to small (2-5 μm) polycrylamide (PA) beads; (b) rabbit anti NADPH-cyt c reductase was immunoadsorbed to the antibody-coated beads; and (c) GF(1+2) was reacted with the beads carrying the two successive layers of antibodies. The beads were then recovered by centrifugation, and were washed, fixed, embedded in agarose, and processed for transmission electromicroscopy. Antireductase- coated beads absorbed 60 percent of the NADPH-cyt c reductase (and comparable fractions of NADH-cyt c reductase and glucose-6-phosphatase) but only 20 percent of the galactosyltransferase activity of the input GF(1+2). Differential vesicle counts showed that approximately 72 percent of the immunoadsorbed vesicles were morphologically recognizable Golgi elements (vesicles with very low density lipoprotein [VLDL] clusters or Golgi cisternae); vesicles with single VLDL and smooth surfaced microsome-like vesicles were too few (approximately 25 percent) to account for the activity. It is concluded that NADPH-cytochrome P(450) reductase is a Golgi membrane enzyme of probably uneven distribution among the elements of the Golgi complex.

Incorporation in vivo of [32P]orthophosphate and [Me-3H]choline into rough microsomal, Golgi, and plasma membranes of rat liver

1977 ◽  
Vol 55 (8) ◽  
pp. 876-885 ◽  
Author(s):  
Patricia L. Chang ◽  
John R. Riordan ◽  
Mario A. Moscarello ◽  
Jennifer M. Sturgess
Keyword(s):  
Endoplasmic Reticulum ◽  
Plasma Membrane ◽  
Rat Liver ◽  
Plasma Membranes ◽  
Specific Activity ◽  
Specific Radioactivity ◽  
Marker Enzyme ◽  
Golgi Membranes ◽  
Endomembrane Flow

To study membrane biogenesis and to test the validity of the endomembrane flow hypothesis, incorporation of 32P and [Me-3H]choline in vivo into membranes of the rat liver was followed. Rough microsomal, Golgi-rich, and plasma membrane fractions were monitored with marker enzyme assays and shown with morphometric analysis to contain 82% rough microsomes, at least 70% Golgi complexes, and 88% plasma membranes, respectively. Membrane subfractions from the rough microsomal and Golgi-rich fractions were prepared by sonic disruption.At 5 to 30 min after 32P injection, the specific radioactivity of phosphatidylcholine was higher in the rough microsomal membranes than in the Golgi membranes. From 1 to 3 h, the specific activity of phosphatidylcholine in Golgi membranes became higher and reached the maximum at about 3 h. Although the plasma membrane had the lowest specific radioactivity throughout 0.25–3 h, it increased rapidly thereafter to attain the highest specific activity at 5 h. Both rough microsomal and plasma membranes reached their maxima at 5 h.The specific radioactivity of [32P]phosphatidylethanolamine in the three membrane fractions was similar to that of [32P]phosphatidylcholine except from 5 to 30 min, when the specific radioactivity of phosphatidylethanolamine in the Golgi membranes was similar to the rough microsomal membranes.At 15 min to 5 h after [Me-3H]choline injection, more than 90% of the radioactivity in all the membranes was acid-precipitable. The specific radioactivities of the acid-precipitated membranes, expressed as dpm per milligram protein, reached the maximum at 3 h. After [Me-3H]choline injection, the specific radioactivity of phosphatidylcholine separated from the lipid extract of the acid-precipitated membranes (dpm per micromole phosphorus) did not differ significantly in the three membrane fractions. The results indicated rapid incorporation of choline into membrane phosphatidylcholine by the rough endoplasmic reticulum, Golgi, and plasma membranes simultaneously.The data with both 32P and [Me-3H]choline precursors did not support the endomembrane flow hypothesis. The Golgi complexes apparently synthesized phosphatidylethanolamine and incorporated choline into phosphatidylcholine as well as the endoplasmic reticulum. The results are discussed with relevance to current hypotheses on the biogenesis and transfer of membrane phospholipids.

scholarly journals On the localisation of d-tubocurarine in rat liver lysosomes in vivo by electron microscopy and subcellular fractionation

1975 ◽  
Vol 289 (3) ◽  
pp. 251-256 ◽  
Author(s):  
Jeanette G. Weitering ◽  
Gerard J. Mulder ◽  
Dirk K. F. Meijer ◽  
Wim Lammers ◽  
Maarten Veenhuis ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Rat Liver ◽  
Subcellular Fractionation ◽  
Liver Lysosomes ◽  
Rat Liver Lysosomes

Poly(A) polymerase activity in ethionine-intoxicated rats: the relative effectiveness of disaggregated and intact polyribosomes as primers for poly(A) polymerase

1980 ◽  
Vol 58 (3) ◽  
pp. 236-242 ◽  
Author(s):  
James Dennis ◽  
Robert Kisilevsky
Keyword(s):  
Rat Liver ◽  
Enzyme Activities ◽  
Relative Effectiveness ◽  
Polymerase Activity ◽  
The State

Ethionine intoxication causes a change in the metabolism of poly(A) sequences on the 3′ OH terminus of mRNA in rat liver in vivo. In an attempt to determine the factors responsible for these changes, nuclear and cytoplasmic poly(A) polymerase activities and the state of the primer were examined in vitro. Requirements for optimal enzyme activities were determined. The nuclear and cytoplasmic enzymes had different K+, Mn2+, and poly(A) primer optima. The levels of nuclear and cytoplasmic poly(A) polymerase activity were shown to decrease following ethionine intoxication. Poly(A)+ RNA isolated from the livers of saline- and ethionine-treated rats served equally well as primers for the cytoplasmic poly(A) polymerase. Disaggregated polysomes were seven times more effective as primers than were intact polysomes. The results suggest that the mRNP particle which is released from polysomes as a result of ethionine intoxication functions better as a poly(A) polymerase primer than does the intact polysome.

Actions ofp-synephrine on hepatic enzyme activities linked to carbohydrate metabolism and ATP levels in vivo and in the perfused rat liver

2017 ◽  
Vol 36 (1) ◽  
pp. 4-12 ◽  
Author(s):  
Marcos Rodrigues Maldonado ◽  
Lívia Bracht ◽  
Anacharis Babeto de Sá-Nakanishi ◽  
Rúbia Carvalho Gomes Corrêa ◽  
Jurandir Fernando Comar ◽  
...  
Keyword(s):  
Carbohydrate Metabolism ◽  
Rat Liver ◽  
Enzyme Activities ◽  
Perfused Rat Liver ◽  
Hepatic Enzyme ◽  
Atp Levels

The in vivo and in vitro effects of alkanols on the morphology and peroxisomal enzyme activities of rat liver

1983 ◽  
Vol 18 ◽  
pp. 64
Author(s):  
C. Rhodes ◽  
C.R. Elcombe ◽  
M.D. Stonard ◽  
M.G. Simpson ◽  
A. Vernall
Keyword(s):  
Rat Liver ◽  
Enzyme Activities ◽  
Peroxisomal Enzyme ◽  
In Vitro Effects
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