Biochemical characterization of the golgi complex of mammalian cells

1974 ◽  
Vol 2 (5-6) ◽  
pp. 737-750 ◽  
Author(s):  
Becca Fleischer ◽  
Fernando Zambrano ◽  
Sidney Fleischer
Keyword(s):  
Golgi Complex ◽  
Mammalian Cells ◽  
Biochemical Characterization

Biochemical Characterization of the Cloned Human 5-HT1A Receptor Expressed in Mammalian Cells

Serotonin ◽  
1990 ◽  
pp. 19-24
Author(s):  
A. Fargin ◽  
J. R. Raymond ◽  
R. J. Lefkowitz ◽  
M. G. Caron
Keyword(s):  
Mammalian Cells ◽  
Biochemical Characterization

Isolation and characterization of the Golgi complex of the protozoan Trypanosoma cruzi

Parasitology ◽  
2001 ◽  
Vol 123 (1) ◽  
pp. 33-43 ◽  
Author(s):  
J. A. MORGADO-DÍAZ ◽  
C. V. NAKAMURA ◽  
O. A. AGRELLOS ◽  
W. B. DIAS ◽  
J. O. PREVIATO ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Trypanosoma Cruzi ◽  
Golgi Complex ◽  
Mammalian Cells ◽  
Sucrose Density Gradient ◽  
Isolation And Characterization ◽  
Golgi Region ◽  
Electron Microscopy Analysis ◽  
Glcnac Transferase

In this study the Golgi complex of the epimastigote forms of Trypanosoma cruzi were isolated and characterized. Using well-controlled sonication to rupture the cells and centrifugation on a discontinuous sucrose density gradient, a highly enriched Golgi fraction was obtained. The Golgi fraction contained most of the β-galactosyltransferase (β-Gal transferase) and UDP-N-acetyl-glucosamine: polypeptide-α-N-acetyl-glucosaminyltransferase (O-α-GlcNAc transferase) activities with minimal contamination of other organelles, as observed by enzymatic assays and electron microscopy analysis. To characterize the Golgi from T. cruzi cells further, it was incubated with a monoclonal antibody against a 58 kDa protein involved in the association of the Golgi complex with microtubules in mammalian cells. Immunofluorescence microscopy showed that the 58 kDa protein is localized in the T. cruzi Golgi region, a result confirmed by high resolution scanning electron microscopy immunocytochemistry. Thus, our results show, for the first time, that the β-Gal transferase, the O-α-GlcNAc transferase and the 58 kDa protein are present in the Golgi complex of T. cruzi and are novel biochemical markers which can be used in the characterization of this organelle in T. cruzi.

Biochemical characterization of the Golgi-complex proteins of Tritrichomonas foetus

1998 ◽  
Vol 84 (9) ◽  
pp. 760-762 ◽  
Author(s):  
José Andrés Morgado Díaz ◽  
Wanderley de Souza
Keyword(s):  
Golgi Complex ◽  
Biochemical Characterization ◽  
Tritrichomonas Foetus

scholarly journals Purification and biochemical characterization of the D6 chemokine receptor

2004 ◽  
Vol 379 (2) ◽  
pp. 263-272 ◽  
Author(s):  
Paul E. BLACKBURN ◽  
Clare V. SIMPSON ◽  
Robert J. B. NIBBS ◽  
Maureen O'HARA ◽  
Rhona BOOTH ◽  
...  
Keyword(s):  
Ligand Binding ◽  
Chemokine Receptor ◽  
Mammalian Cells ◽  
Biochemical Characterization ◽  
Mutational Analysis ◽  
Three Dimensional ◽  
Dimensional Structure ◽  
Affinity Column ◽  
Transfected Cells

There is much interest in chemokine receptors as therapeutic targets in diseases such as AIDS, autoimmune and inflammatory disorders, and cancer. Hampering such studies is the lack of accurate three-dimensional structural models of these molecules. The CC-chemokine receptor D6 is expressed at exceptionally high levels in heterologous transfectants. Here we report the purification and biochemical characterization of milligram quantities of D6 protein from relatively small cultures of transfected mammalian cells. Importantly, purified D6 retains full functional activity, shown by displaceable binding of 125I-labelled MIP-1β (macrophage inflammatory protein-1β) and by complete binding of the receptor to a MIP-1α affinity column. In addition, we show that D6 is decorated on the N-terminus by N-linked glycosylation. Mutational analysis reveals that this glycosylation is dispensable for ligand binding and high expression in transfected cells. Metabolic labelling has revealed the receptor to also be sulphated and phosphorylated. Phosphorylation is ligand independent and is not enhanced by ligand binding and internalization, suggesting similarities with the viral chemokine receptor homologue US28. Like US28, an analysis of the full cellular complement of D6 in transfected cells indicates that >80% is found associated with intracellular vesicular structures. This may account for the high quantities of D6 that can be synthesized in these cells. These unusual properties of D6, and the biochemical characterization described here, leads the way towards work aimed at generating the three-dimensional structure of this seven-transmembrane-spanning receptor.

Interaction proteomics: characterization of protein complexes using tandem affinity purification–mass spectrometry

2010 ◽  
Vol 38 (4) ◽  
pp. 883-887 ◽  
Author(s):  
Pamela Völkel ◽  
Perrine Le Faou ◽  
Pierre-Olivier Angrand
Keyword(s):  
Protein Complex ◽  
Mammalian Cells ◽  
Protein Identification ◽  
Biochemical Characterization ◽  
Protein Complexes ◽  
Affinity Purification ◽  
Protein A ◽  
High Sensitivity ◽  
Tandem Affinity Purification

Most cellular processes are carried out by a multitude of proteins that assemble into multimeric complexes. Thus a precise understanding of the biological pathways that control cellular events relies on the identification and on the biochemical characterization of the proteins involved in such multimeric assemblies. Advances in MS have made possible the identification of multisubunit protein complexes isolated from cell lysates with high sensitivity and accuracy, whereas the TAP (tandem affinity purification) methodology efficiently isolates native protein complexes from cells for proteomics analysis. TAP is a generic method based on the sequential utilization of two affinity tags to purify protein assemblies. During the first purification step, the Protein A moiety of the TAP tag is bound to IgG beads, and protein components associated with the TAP-tagged protein are retrieved by TEV (tobacco etch virus) protease cleavage. This enzyme is a sequence-specific protease cleaving a seven-amino-acid recognition site located between the first and second tags. In the second affinity step, the protein complex is immobilized to calmodulin-coated beads via the CBP (calmodulin-binding peptide) of the TAP tag. The CBP–calmodulin interaction is calcium-dependent and calcium-chelating agents are used in the second elution step to release the final protein complex preparation used for protein identification by MS. The TAP–MS approach has proven to efficiently permit the characterization of protein complexes from bacteria, yeast and mammalian cells, as well as from multicellular organisms such as Caenorhabditis elegans, Drosophila and mice.

Plant Pathology Diagnosed by X-Ray Microanalysis

1987 ◽  
Vol 45 ◽  
pp. 974-975
Author(s):  
J. H. Resau ◽  
N. Howell ◽  
S. H. Chang
Keyword(s):  
Electron Microscopy ◽  
Plant Pathology ◽  
Biochemical Characterization ◽  
Nutrient Deficiency ◽  
Green Leaf ◽  
Parasitic Infestation ◽  
X Ray

Spinach grown in Texas developed “yellow spotting” on the peripheral portions of the leaves. The exact cause of the discoloration could not be determined as there was no evidence of viral or parasitic infestation of the plants and biochemical characterization of the plants did not indicate any significant differences between the yellow and green leaf portions of the spinach. The present study was undertaken using electron microscopy (EM) to determine if a micro-nutrient deficiency was the cause for the discoloration.Green leaf spinach was collected from the field and sent by express mail to the EM laboratory. The yellow and equivalent green portions of the leaves were isolated and dried in a Denton evaporator at 10-5 Torr for 24 hrs. The leaf specimens were then examined using a JEOL 100 CX analytical microscope. TEM specimens were prepared according to the methods of Trump et al.

Interaction effect of rootstocks on gas exchange parameters, biochemical changes and nutrientstatus in Sauvignon Blanc winegrapes

2014 ◽  
Vol 3 (3) ◽  
pp. 218-225
Author(s):  
R. G. Somkuwar ◽  
M. A. Bhange ◽  
A. K. Upadhyay ◽  
S. D. Ramteke
Keyword(s):  
Biochemical Characterization ◽  
Reducing Sugar ◽  
Wine Grape ◽  
Gas Exchange Parameters ◽  
Food Material ◽  
Total Carbohydrates ◽  
Salt Creek ◽  
Grape Varieties ◽  
Photosynthetic Activities

SauvignonBlanc wine grape was characterized for their various morphological, physiological and biochemical parameters grafted on different rootstocks. Significant differences were recorded for all the parameters studied. The studies on vegetative parameters revealed that the rootstock influences the vegetative growth thereby increasing the photosynthetic activities of a vine. The highest photosynthesis rate was recorded in 140-Ru grafted vine followed by Fercal whereas the lowest in Salt Creek rootstock grafted vines.The rootstock influenced the changes in biochemical constituents in the grafted vine thereby helping the plant to store enough food material. Significant differences were recorded for total carbohydrates, proteins, total phenols and reducing sugar. The vines grafted on1103-Pshowed highest carbohydrates and starch followed by 140-Ru,while the least amount of carbohydrates were recorded in 110-R and Salt Creek grafted vines respectively.Among the different rootstock graft combinations, Fercal showed highest amount of reducing sugar, proteins and phenols, followed by 1103-P and SO4, however, the lowest amount of reducing sugar, proteins and phenols were recorded with 110-R grafted vines.The vines grafted on different rootstocks showed changes in nutrient uptake. Considering this, the physico-biochemical characterization of grafted vine may help to identify particularrootstocks combination that could influence a desired trait in commercial wine grape varieties after grafting.

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