Abstract
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. Key words: cell disruption, purification, prokaryote, eukaryote, functional unit.
Highlights
The tree of life is made up of three distinct domains: eukaryotes, bacteria, and archaea (Forterre, 2001)
This paper examines the structure of some compartments in microbial cells and the method of isolation and purification of these structures
To obtain a purified plasma membrane fraction from epimastigotes of T. cruzi, cells previously swollen by treatment with Triton X-100 were disrupted using a Dounce-type homogenizer followed by consecutive steps of differential centrifugation and its purity was evaluated by electron microscopy and by testing for enzyme markers
Summary
The tree of life is made up of three distinct domains: eukaryotes, bacteria, and archaea (Forterre, 2001). To obtain a purified plasma membrane fraction from epimastigotes of T. cruzi, cells previously swollen by treatment with Triton X-100 were disrupted using a Dounce-type homogenizer followed by consecutive steps of differential centrifugation and its purity was evaluated by electron microscopy and by testing for enzyme markers. Protein can be extracted by a few methods such as detergent lysis, shearing force, treatment with low ionic salt (salting out), and rapid changes in pressure, which aimed to weaken and break the membranes surrounding the cell to allow proteins to escape (Watson et al, 2004) Mechanical disruption techniques, such as French Press or glass beads are usually used to remove the cell wall, followed by detergent based extraction of total protein The sample was eluted with a discontinuous gradient with a salt-containing buffer (MoraMontes et al, 2010)
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