In SDS-PAGE, the relative mobility and the log molecular weight as given by

.......………………………………………Eq (22.1)

Molecular weight of a protein can be determined by plotting relative migration Rf with the log molecular weight of standard protein.

…………Eq (22.2)

Figure 22.4: SDS-PAGE Profile of a typical bacterial lysate.

 

2. Native PAGE: SDS-PAGE discussed in the previous lecture is using anionic detergent sodium dodecyl sulfate and β-mercaptoethanol to give equal charge to all protein and breaks the disulphide linkage. As a result, the 3-D structure of the protein is destroyed and it migrate as per their subunit molecular weight. In the native PAGE, sample is prepared in the loading dye does not contains detergent or denaturating agent and as a result sample runs on the basis of charge/mass. In native PAGE, the 3-D conformation as well as activity of the protein remains unaffected.

3. Urea PAGE: In this method, insoluable protein is dissolved in Urea and samples separate based on their charge/subunit mass. A gradient Ura PAGE is used to monitor the folding states of a protein.