Cell fractionation of parasitic protozoa: a review

Souza, Wanderley de; Cunha-e-Silva, Narcisa L.

doi:10.1590/s0074-02762003000200001

Cited by 16 publications

(11 citation statements)

References 151 publications

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“…Several techniques utilized included ultrasound disintegration, homogenization in bead mill, application of chemicals of various types, and osmotic shock. Usually, it is necessary to monitor the process by phase contrast microscopy in order to avoid rupture of the compartments or disorganization of cytoplasmic structures (de Souza and Cunha da Silva, 2003).…”

Section: Cell Disruptionmentioning

confidence: 99%

“…Usually differential centrifugation does not yield very pure material. Therefore, it is necessary to continue the isolation procedure by using density-gradient centrifugation, a procedure where the compartments and structures are separated by sedimentation through a gradient of a dense solution such as sucrose, Metrizamide, Percoll (de Souza and Cunha da Silva, 2003).…”

Section: Cell Fractionationmentioning

confidence: 99%

“…After centrifugation, six fractions of equal volume were collected from the top to bottom. The third fraction was a highly purified para-flagellar rod fraction (de Souza and Cunha-e-Silva, 2003). Cunha-eSilva et al (1989) managed to get a highly purified flagellar membrane fraction from H. megaseliae and T. cruzi epimastigotes by combining detergent treatment on ice, vortexing and separation on a sucrose equilibrium gradient.…”

Section: Flagellummentioning

confidence: 99%

“…The protozoa contain unique cytoplasmic structures and organelles (de-Souza and da-Cunha-e-Silva, 2003). These compartments are similar to those of other eukaryotic cells.…”

Section: Introductionmentioning

confidence: 99%

“…Eukaryotes include animals, plants, fungi, and protozoa. The protozoa group includes an enormous number of species, comprising some agents of human and animal diseases such as malaria, and leishmaniasis (de Souza and da Cunha-e-Silva, 2003). Most fungi are filamentous, many grow as unicellular yeasts (Saccharomyces cerevisiae) while some for example chytridomycetes grow as individual rounded cells or dichotomous branched chains of cells with root-like rhizoids for attachment to a nutrient resource (Kavanagh, 2011).…”

Section: Introductionmentioning

confidence: 99%

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How to obtain the organelles of prokaryotic and microbial eukaryotic cells

Aderiye¹,

Oluwole²

2014

J. Cell Anim. Biol.

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An organelle is a specialized functional subunit within cells carrying out specific functions. These compartments which may or may not be enclosed in a lipid bilayer are found in microorganisms. While those found in eukaryotic cells are usually enclosed in lipid bilayer, those in prokaryotes don't. All microbes have compartments common to them like the nucleic acids, protein, ribosomes as well as unique intracellular structures found only in microbial subgroups. Such compartments include the mitochondria, endoplasmic reticulum, golgi apparatus amongst others unique to all eukaryotic cells only. Prokaryotes contain some micro-compartments unique to them including the carboxysomes, lipid bodies, polyhydroxybutyrate granules. The right choice of cell disruption methods that limit damage to the compartments is important in achieving successful compartment isolation and purification. Commonly applied methods include sonication, enzymatic lysis, detergent lysis, cavitation amongst others depending on the type of cells involved. Fractionation is the commonly utilized method for isolation and purification of organelles, utilizing ultracentrifugation and techniques that exploits size, density and surface charge variations of protoplasmic content. Such techniques include gradient centrifugation methods, use of beads, affinity purification chromatography methods and electrophoresis. Here, we review the compartments in microbial cells and the techniques employed to isolate and purify these intracellular components.

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Section: Cell Disruptionmentioning

confidence: 99%

Section: Cell Fractionationmentioning

confidence: 99%

Section: Flagellummentioning

confidence: 99%

“…The protozoa contain unique cytoplasmic structures and organelles (de-Souza and da-Cunha-e-Silva, 2003). These compartments are similar to those of other eukaryotic cells.…”

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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How to obtain the organelles of prokaryotic and microbial eukaryotic cells

Aderiye¹,

Oluwole²

2014

J. Cell Anim. Biol.

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Plant organelle proteomics: Collaborating for optimal cell function

Agrawal¹,

Bourguignon

Rolland

et al. 2010

Mass Spectrometry Reviews

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Organelle proteomics describes the study of proteins present in organelle at a particular instance during the whole period of their life cycle in a cell. Organelles are specialized membrane bound structures within a cell that function by interacting with cytosolic and luminal soluble proteins making the protein composition of each organelle dynamic. Depending on organism, the total number of organelles within a cell varies, indicating their evolution with respect to protein number and function. For example, one of the striking differences between plant and animal cells is the plastids in plants. Organelles have their own proteins, and few organelles like mitochondria and chloroplast have their own genome to synthesize proteins for specific function and also require nuclear-encoded proteins. Enormous work has been performed on animal organelle proteomics. However, plant organelle proteomics has seen limited work mainly due to: (i) inter-plant and inter-tissue complexity, (ii) difficulties in isolation of subcellular compartments, and (iii) their enrichment and purity. Despite these concerns, the field of organelle proteomics is growing in plants, such as Arabidopsis, rice and maize. The available data are beginning to help better understand organelles and their distinct and/or overlapping functions in different plant tissues, organs or cell types, and more importantly, how protein components of organelles behave during development and with surrounding environments. Studies on organelles have provided a few good reviews, but none of them are comprehensive. Here, we present a comprehensive review on plant organelle proteomics starting from the significance of organelle in cells, to organelle isolation, to protein identification and to biology and beyond. To put together such a systematic, in-depth review and to translate acquired knowledge in a proper and adequate form, we join minds to provide discussion and viewpoints on the collaborative nature of organelles in cell, their proper function and evolution.

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Subcellular proteomics of Trypanosoma cruzi reservosomes

et al. 2009

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Reservosomes are the endpoint of the endocytic pathway in Trypanosoma cruzi epimastigotes. These organelles have the particular ability to concentrate proteins and lipids obtained from medium together with the main proteolytic enzymes originated from the secretory pathway, being at the same time a storage organelle and the main site of protein degradation. Subcellular proteomics have been extensively used for profiling organelles in different cell types. Here, we combine cell fractionation and liquid chromatography-tandem mass spectrometry (LC-MS/MS) analysis to identify reservosome-resident proteins. Starting from a purified reservosome fraction, we established a protocol to isolate reservosome membranes. Transmission electron microscopy was applied to confirm the purity of the fractions. To achieve a better coverage of identified proteins we analyzed the fractions separately and combined the results. LC-MS/MS analysis identified in total 709 T. cruzi-specific proteins; of these, 456 had predicted function and 253 were classified as hypothetical proteins. We could confirm the presence of most of the proteins validated by previous work and identify new proteins from different classes such as enzymes, proton pumps, transport proteins and others. The definition of the reservosome protein profile is a good tool to assess their molecular signature, identify molecular markers, and understand their relationship with different organelles.

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Cell fractionation of parasitic protozoa: a review

Cited by 16 publications

References 151 publications

How to obtain the organelles of prokaryotic and microbial eukaryotic cells

How to obtain the organelles of prokaryotic and microbial eukaryotic cells

Plant organelle proteomics: Collaborating for optimal cell function

Subcellular proteomics of Trypanosoma cruzi reservosomes

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