Be introduced to the world of analytical techniques in the purification of biomolecules in this free course. You will explore the fact that chromatography is a technique that allows scientists to separate molecules of interest from a crude mixture and discover which effect this separation exploits. Find out which type of chromatography leads to the matric having a negatively charged functional group with an affinity towards positively charged molecules. You will be introduced to the concept of ‘hydrophobic interaction chromatography’ which exploits the ability of strong interactions between the hydrophobic group attached to the matrix and hydrophobic patches present on an analyte such as a protein. Find out what happens to the water molecules shielding the protein side chains in the presence of a high amount of salt, and discover what the addition of a low amount of salt to the protein solution results in. Demonstrations, problems, examples and assessment questions in this instructor-led video-based course are designed to provide you with a comprehensive foundation in this aspect of biology.
The course kicks off with the definition of gel filtration chromatography (also known as size-exclusion chromatography or molecular-sieve chromatography) and the different types of problems that one is likely to have after packing the column. The choice of the column will be discussed, depending on the range of molecular weight and the pressure limit of the operating equipment. You will explore the fact that the molecular weight and size of a protein is related to the shape of the molecule and the relationship between its molecular weight and the radius of gyration. You will discover the fact that the native molecular weight determination by gel filtration in conjugation with the SDS-PAGE can be used to determine the oligomeric status of the protein. Go on to study preparing gel filtration samples in the mobile phase and how the binding of DNA to protein induces certain conformational changes.
Next, you will learn that affinity chromatography works on the principle of mutual recognition forces between a ligand and a receptor. You will gain insight into the fact that in the bio-affinity chromatography method, biomolecules are used as a receptor present on the matrix and it exploits the biological phenomenon such as the antibody-antigen. You will then discover that Glutathione S-transferase (GST) utilizes glutathione as a substrate to catalyze conjugation reactions for xenobiotic detoxification purposes. This GST fusion protein is produced by the recombining of the protein of interest with the GST sequence present in the expression vector. You will be taught that the biotinylation of antibodies allows the immobilization of antibodies in the correct orientation on the streptomycin coated glass beads. You will discover that an affinity column can also be used as a tool to study or isolate an interacting partner of a particular protein. Finally, you will study gel filtration chromatography, receptor and ligand interactions, bio-affinity chromatography, and metal-affinity chromatography. This course will be of interest to biotechnology students, or those professionals in experimental biotechnology.
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View All Learning Outcomes View Less All Alison courses are free to enrol study and complete. To successfully complete this course and become an Alison Graduate, you need to achieve 80% or higher in each course assessment. Once you have completed this course, you have the option to acquire an official Diploma, which is a great way to share your achievement with the world.
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