SDS (sodium dodecyl sulfate) and DTT (dithiothreitol) are chemicals commonly used in molecular biology, particularly in techniques like SDS-PAGE (sodium dodecyl sulfate polyacrylamide gel electrophoresis) to prepare proteins for analysis.
SDS: Sodium Dodecyl Sulfate
- What it is: SDS is an anionic detergent (surfactant).
- How it works:
- Denaturation: It disrupts the non-covalent bonds in proteins, causing them to unfold and lose their native structure.
- Charging: SDS binds to proteins and coats them with a negative charge. This uniform negative charge masks the protein's intrinsic charge, ensuring that proteins migrate through an electrophoresis gel based on their size, not their charge.
- Solubilization: SDS helps to solubilize proteins, preventing them from aggregating during electrophoresis.
- Why it's important: SDS ensures that proteins separate solely based on their molecular weight during SDS-PAGE. Without SDS, proteins would separate based on both size and charge, leading to inaccurate results.
DTT: Dithiothreitol
- What it is: DTT is a reducing agent.
- How it works:
- Reduces Disulfide Bonds: DTT breaks disulfide bonds (S-S bonds) that form between cysteine amino acids in a protein. These bonds contribute to a protein's tertiary and quaternary structure. Breaking them allows for complete protein denaturation.
- Prevents Oxidation: DTT also helps prevent oxidation of proteins by scavenging free radicals.
- Why it's important: Disulfide bonds can hinder proper protein separation during electrophoresis. By reducing these bonds, DTT ensures that proteins are fully unfolded and can migrate freely through the gel matrix. It is often used in conjunction with SDS for optimal denaturation. For instance, it can be used to denature CD38 on red blood cells.
In summary, SDS denatures proteins and provides a uniform negative charge, while DTT reduces disulfide bonds to ensure complete protein unfolding. They work synergistically to prepare proteins for accurate size-based separation during SDS-PAGE.
