DENATURATION OF PROTEINS
The experiment was done to be able to understand how various denaturants such as HCl and NaOH affects proteins. It was observed that different denaturants act upon or denature protein differently. This was determined using the principle of viscometry. An Ostwald viscometer was used to measure the viscosity of the prepared native, blank, denatured native and blank with denaturant solutions. The time required for the said solutions to pass through the viscometer were recorded and their viscosities were then calculated using the data. Experimentally, urea was found to be the best out of the six denaturants that were used.
Results and Discussion
Proteins differ from each other by means of the amino acids that it contains. Each protein has its own biological function based on the way its components are arranged, thus, a change in the conformation of these components will cause a drastic change in its function. And when this happens, the protein is said to be denatured. Denaturation refers to the state wherein there is a disruption in the original conformation of the protein – that is the native state of the protein. Only the primary of structure of the protein remains unchanged during denaturation while the secondary, tertiary and quaternary structures are all affected. Denaturation is brought about by various kinds of physical and chemical means; this includes changes in pH and temperature, or the addition of organic compounds, detergents, metal ions, and alkaloid reagents. The bonds that stabilize a protein’s tertiary structure, its salt linkage, hydrogen bonds, disulfide linkage and hydrophobic interactions, once broken, disrupt the protein’s secondary and tertiary structure. Aside from the change in its biological function, other effects of denaturation include the decrease in the solubility of the protein and increase in its isoelectric pH.
The change in pH causes denaturation in a way that when the pH of the protein is low (that is, it is acidic) the molecules become positively charged and as a result, the protein uncoils because of the repulsion between the positively charged molecules. Denaturation may also results from the increase in temperature, bringing about an increase in the kinetic energy of the protein. It is known that at high temperatures, molecules moves faster compared to when the temperature is low; thus, high temperature causes an increase in the kinetic energy of the molecules which in turn causes vibrations that disrupt the relatively weak hydrogen bonds and hydrophobic bonds of the protein.
Strong acids denature proteins by altering the state of ionization of the carboxyl side chains, hence disrupting salt linkages. Organic compounds such as ethanol causes denaturation in such a way that it causes a disruption in the hydrogen bonds of the protein molecule because of its capability of forming H-bonds. The disinfecting power of alcohols could be accounted to its capability to denature the proteins of bacteria thereby killing them. The metal ions like Pb2+, Ag+, Hg2+ ions and alkaloid reagents which affects the salt bridges and disulfide linkages of proteins are also considered as denaturants. Detergents on the other hand affect the hydrophobic side chains of the protein molecule.
Viscosity is the rate of flow of a liquid or the measure of its resistance to flow. It is dependent on the magnitude of the intermolecular forces of the molecule; the stronger the intermolecular force, the more viscous the liquid is. Viscosity is also dependent on the size of the molecule.
In this experiment, the effect of denaturation on the viscosity of albumin and casein were investigated; the two proteins were subjected to six different denaturants. The viscosity of the blank solution, 0.1 M NaOH for casein and 0.9% NaCl for albumin, was measured using an Ostwald viscometer; the viscosity of the native, blank with denaturant and native with denaturant were...
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