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Cell Organelles: Cell Fractionation & Assay Essay

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Cell Organelles: Cell fractionation & Assay Introduction: Cells require many cellular processes to function properly. Eukaryotic cells organize some of their cellular processes in membrane-bound vesicles called organelles. Each organelle facilitates cellular processes by combining related compounds in an enclosed space and separating interfering reactions (Ardell). One example of this is the mitochondria. The mitochondria are a compartment of the cell where the production of energy is catalyzed through several steps. It is here that molecules of glucose enter from other parts of the cell which then go through glycolysis. The products of glycolysis, two pyruvate molecules, then undergo the citric acid cycle which involves a series of oxidation reactions to yield 2 molecules of CO2, 3 molecules of NADH, 1 molecule of GTP and 1 molecules of FADH2 for each turn of the cycle (Alberts). In this lab, we focus on the sixth step in the reaction where succinate is oxidized to fumarate with the help of the enzyme succinate dehydrogenase and forms FADH2 (Alberts). The enzyme used in this step is unique to the mitochondria and therefore can be used as a marker for the presence of mitochondrial contamination in a sample of cell fraction. We can use cellular fractionation to separate the cellular components and test them for the presence of certain enzymes or chemicals (Hames). Cellular fractionation involves two steps. Firsty, we must lyse the cells from a homogenous cell population to obtain a homogenate of subcellular components. Since different cellular components vary in size and weight, we can centrifuge them at different speeds and durations to obtain pellets containing these cell fractions (Ardell). We can use the cell fractions obtained to perform an enzyme assay in order to determine the contamination in our fractions. In this lab, we focus on the enzyme activity of succinate dehydrogenase to detect the contamination of mitochondrial material in different cell...

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