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In cell biology, microsomes are heterogeneous vesicle-like artifacts (~20-200 nm diameter) re-formed from pieces of the endoplasmic reticulum (ER) when eukaryotic cells are broken-up in the laboratory; microsomes are not present in healthy, living cells.
Rough (containing ) and smooth (without ribosomes) microsomes are made from the endoplasmic reticulum through cell disruption. These microsomes have an inside that is exactly the same as the endoplasmic reticulum lumen. Both forms of microsomes can be purified by a process known as equilibrium density centrifugation. Rough and smooth microsomes do differ in their proteins and rough microsomes have shown occurrence of translation and translocation at the same time besides certain exceptions from proteins in yeast.
Protein extrusion into a microsome can be described by multiple factors. A protein has been extruded if it is resistant to proteases, is not resistant to proteases when detergents are present, or is glycosylated by enzymes residing in the microsomes. Additionally, another sign that a protein has been extruded is signal peptidase cleaving off the N-terminal signal peptide inside the microsome that may cause the protein to be smaller in size.
Other experiments have shown that microsomes have to be introduced before about the first 70 are translated for the secretory protein to go into the microsomal lumen. At this point, 40 amino acids are sticking out from the ribosome and the 30 amino acids after that are in the ribosomal channel. Cotranslational translocation explains that transport into the endoplasmic reticulum lumen of secretory proteins starts with the protein still bound to the ribosomes and not completely synthesized.Lodish, H. F. et al. (2008). Molecular Cell Biology. W.H. Freeman. Microsomes can be concentrated and separated from other cellular debris by differential centrifugation. Unbroken cells, Cell nucleus, and mitochondria sediment out at 10,000 g (where g is the Earth's gravitational acceleration), whereas soluble enzymes and fragmented ER, which contains cytochrome P450 (CYP), remain in solution. At 100,000 g, achieved by faster centrifuge rotation, ER sediments out of solution as a pellet but the soluble enzymes remain in the supernatant. In this way, cytochrome P450 in microsomes is concentrated and isolated. Microsomes have a reddish-brown color, due to the presence of the heme. Because of the need for a multi-part protein-system, microsomes are necessary to analyze the Metabolism of CYPs. These CYPs are highly abundant in livers of rats, mice and humans, but present in all other organs and organisms as well.
To get microsomes containing a specific CYP or for high amounts of active enzyme, microsomes are prepared from Sf9 insect cells or in yeast via heterologous expression. Alternatively expression in Escherichia coli of whole or truncated proteins can also be performed. Therefore, microsomes are a valuable tool for investigating the metabolism of compounds (enzyme inhibition, clearance and metabolite identification) and for examining drug-drug interactions by in vitro-research. Researchers often select microsome lots based on the enzyme activity level of specific CYPs. Some lots are available to study specific populations (for example, lung microsomes from smokers or non-smokers) or divided into classifications to meet target CYP activity levels for inhibition and Metabolomics.
Microsomes are used to mimic the activity of the endoplasmic reticulum in a test tube and conduct experiments that require protein synthesis on a membrane. They provide a way for scientists to figure out how proteins are being made on the ER in a cell by reconstituting the process in a test tube.
Keefer et al. looked into how human liver microsomes and human are used to study metabolic stability and inhibition for in vitro systems. Going into their similarities and differences can shine light on the mechanisms of metabolism, passive permeability, and transporters. It was shown that passive permeability is important in metabolism and enzyme inhibition in human hepatocytes. Also, P-gp efflux has a smaller role in this same area. Also, liver microsomes are more predictive than hepatocytes of in vivo clearance when they give higher intrinsic clearance than the hepatocytes.
Wang et al. explored drug metabolism in vitro using human liver microsomes and human liver S9 fractions. The study found significant differences between human liver microsomes and human liver S9 fractions in drug-metabolizing enzyme and transporter protein concentrations. The protein-protein correlations of these drug-metabolizing enzymes and transporters was determined relating to the two hepatic preparations.
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