The purification continues with a cation exchange step in which the positively charged cytochrome C protein is separated from negatively charged DNA and other proteins. The cation exchange eluate (volume of solution collected) had a total volume of 42.0 mL and a 1.0 mL aliquot was set aside for further analysis. The following data was obtained from the 1.0 mL aliquot to quantify the protein amount and purity:
Enzyme kinetics
In biochemistry, enzymes are proteins that act as biological catalysts. Catalysis is the addition of a catalyst to a chemical reaction to speed up the pace of the reaction. Catalysis can be categorized as either homogeneous or heterogeneous, depending on whether the catalysts are distributed in the same phase as that of the reactants. Enzymes are an essential part of the cell because, without them, many organic processes would slow down and thus will affect the processes that are important for cell survival and sustenance.
Regulation of Enzymes
A substance that acts as a catalyst to regulate the reaction rate in the living organism's metabolic pathways without itself getting altered is an enzyme. Most of the biological reactions and metabolic pathways in the living systems are carried out by enzymes. They are specific for their works and work in particular conditions. It maintains the best possible rate of reaction in the most stable state. The enzymes have distinct properties as they can proceed with the reaction in any direction, their particular binding sites, pH specificity, temperature specificity required in very few amounts.
The purification continues with a cation exchange step in which the positively charged cytochrome C protein is separated from negatively charged DNA and other proteins. The cation exchange eluate (volume of solution collected) had a total volume of 42.0 mL and a 1.0 mL aliquot was set aside for further analysis. The following data was obtained from the 1.0 mL aliquot to quantify the protein amount and purity:
- The absorbance at 410 nm of the aliquot was diluted 5-fold was 0.474 (1 cm pathlength).
- The absorbance at 595 nm from a 1.0 mL Bradford Assay solution that was diluted by 100-fold from the aliquot was 0.195 (1 cm pathlength).
Using the information given,
- Calculate the total protein amount in mg from the absorbance at 595 nm.
- Calculate the cytochrome C amount in mg from the absorbance at 410 nm using Beer’s Law.
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