Question:
What role do detergents play in protein solubilization?
The Protein Man Says:
Detergents are commonly defined as a class of molecules that exhibit an amphipathic structure. All detergents have a hydrophilic (water-loving) polar head and a hydrophobic (water-fearing) non-polar tail. Due to their unique structure, they have the ability to form or disrupt hydrophilic-hydrophobic interactions in most biological samples. Aside from its role in protein solubilization, detergents also play an important function in the following procedures:
-
cell lysis
-
protein crystallization
-
electrophoresis
-
prevention of non-specific binding in affinity purification and immunoassay procedures
Detergents and Protein Solubilization – Understanding the Process
In aqueous solutions, the detergent's polar head interacts with the hydrogen bonds of the water molecules while the non-polar tail ends aggregate to form highly organized spherical structures known as micelles. The concentration at which micelles begin to form (known as the Critical Micelle Concentration or CMC) is of vital importance since it provides the researcher the precise amount of detergent that should be used to allow for complete protein solubilization.
So, how do detergents release or solubilize proteins? Here's how.
Most lipids and proteins are embedded in biological membranes which consist of amphipathic phospholipid bi-layers which have almost the same structure as the detergent micelles. While these proteins are not soluble in aqueous solutions, they can be released from the lipid bi-layer by using an appropriate detergent.
Upon the introduction of moderate amounts of biological detergents (less than the detergent's CMC) into the aqueous solution, the detergent molecules begin to disrupt the biological membrane where the proteins are embedded. However, at concentrations equal to or higher than the detergent's CMC, the lipid bi-layer breaks apart and the hydrophobic end of the detergent micelle binds with the hydrophobic end of the protein to prevent them from aggregating.
Types of Detergents
There are several types of detergents that can be used for your experimentations. These include the following:
-
Non-ionic detergents. These are the mild, non-denaturing detergents that researchers prefer to use in breaking lipid-lipid and lipid-protein interactions. They facilitate protein solubilization without compromising the native sub-unit structure or any of its functions.
-
Ionic detergents. Anionic and cationic detergents are classified as biologically harsh detergents and are commonly used in DNA purification. They can cause complete disruption of the cellular structure and may completely denature proteins during gel electrophoresis. SDS, CTAB and deoxycholate are all included in this group.
-
Zwitterionic detergents. Detergents belonging to this group exhibit the characteristics of both ionic and non-ionic detergents. As such, they can protect the native state of the protein without changing the native charge of the protein molecules. Zwitterionic detergents are used for isoelectric focusing, 2D electrophoresis and ion-exchange chromatography.
