N-Acetyl-Β-Hexosaminidase Lab Report

The hypothesis of the experiment was that the molecular weight of N-acetyl-β-D-hexosaminidase would be around 28,000 kDa but to determine the molecular weight the concept of SDS-PAGE had to be understood. The movement of any charged particle through an electric field is determined by its net charge, its molecular radius and the magnitude of the applied field (7). Being that natively folded proteins are not molecular weight dependent, the proteins would be traveling at different speeds in an electric field. In order to separate the proteins based on their molecular weight, the tertiary structure of the protein needs to be destroyed making the protein a linear molecule. Sodium dodecyl sulfate (SDS) plays an important role in this mechanism. Sodium dodecyl sulfate is an amphipathic detergent whose role is to break down the tertiary structure of a protein making it a linear molecule and also masking the native charge of the protein with a negative charge. By using SDS the protein involved in this experiment allowed the different volumes of the protein to migrate down the electrophoresis gel at the same time. SDS is also present throughout the gel to make sure the proteins are linearized and their charges are masked throughout the run. The gel run is conducted …show more content…
The Rf value for N-acetyl-β-D-hexosaminidase was calculated using the distance of the migration of the protein divided by the total migration of the solvent resulting in a Rf value of 0.3. The linear equation generated from the graph (Figure 2) was used to determine the log (MW), which was determined to be 4.9. The antilog of the log (MW) was used to determine to be 79,432