Why would the experimental value of molecular weight of catalase be much lower than literature values as determined by an SDS PAGE analysis? (The experimental value that I got was 58,000 daltons, which is much lower than it should be)
The experimental value of the molecular weight of catalase being much lower than literature values as determined by an SDS-PAGE analysis can be due to several factors. Here are some possible explanations:
1. Protein degradation: Proteins can undergo degradation during the experimental procedure, leading to the formation of smaller fragments. This can happen due to enzymatic activity or denaturation during sample preparation, storage, or analysis. It is important to handle the enzyme carefully, avoid excessive heat, and use appropriate buffers to minimize degradation.
2. Denaturation: Catalase can undergo denaturation under certain conditions such as extreme pH, high temperatures, or exposure to harsh chemicals. Denaturation can disrupt the protein's tertiary and quaternary structure, causing it to migrate differently on an SDS-PAGE gel. Make sure you are using the appropriate conditions to preserve the protein's native structure.
3. Sample preparation: Proper sample preparation is crucial for accurate molecular weight determination. If there were errors in sample preparation, such as incorrect sample concentration, improper sample loading, or incomplete reduction and denaturation processes, it can affect the migration pattern on the gel.
4. SDS-PAGE limitations: SDS-PAGE can provide approximate molecular weight estimation, but it is not always precise. Depending on the protein's shape, charge, and interaction with the SDS detergent, the migration rate on the gel might not directly correlate with the molecular weight. Different proteins can migrate differently on the gel even if they have similar molecular weights.
If you obtained a significantly lower molecular weight value, it would be advisable to perform additional analyses to confirm the results. Some other techniques that can provide more accurate molecular weight determination include size exclusion chromatography, mass spectrometry, or analytical ultracentrifugation.