Influence of Ionic Strength, pH, and SDS Concentration on Subunit Analysis of Phycoerythrins by SDS-PAGE

Abstract

Sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) is often used for subunit analysis of proteins, but it is not efficient to make the alpha- and beta-subunits of phycoerythrins separated by normal SDS-PAGE. In this research, subunit components and subunit molecular weights of four purified phycoerythrins were analyzed by SDS-PAGE. Four factors including Tris concentration, pH, ammonium persulfate (APS), and SDS concentration were studied for their effects on SDS-PAGE of phycoerythrins. It showed that these factors can influence the separation of alpha- and beta-subunits, electrophoresis effect of gamma-subunits, apparent molecular weights of subunits, and mobility of marker proteins. The alpha- and beta-subunits separated better in the case of lower SDS concentration, lower Tris concentration, higher pH, and/or lower APS addition in separating gels. The molecular weights of alpha- and beta-subunits increased when Tris concentration increased in a certain range. It can be concluded that factors critical to subunit analysis by SDS-PAGE are SDS concentration and ionic strength, both of which are related to critical micelle concentration of SDS and ratio of SDS monomer to micelle in SDS-PAGE system. The ratio is postulated to influence SDS-PAGE by influencing the amount of SDS bound to polypeptides and shapes of polypeptide-SDS complexes.

Effect of pH on physicochemical properties of oil bodies from different oil crops

“…The protein bands of PNOBs weakened at pH 2-4 and pH 10-12, and the protein bands witness the most prominent reduction at pH 2 ( Figure 2S b). Acidic and alkaline environments can all lead to the reduction of some oil mosaic proteins and storage proteins which is in agreement with Zhao et al (2010). Since the pH can affect the surface charge properties of OBs and the interaction between phospholipid and protein, it may lead to the rearrangement of the surface proteins of OBs (Mohamed et al 2005).…”

“…The surface proteins of SBOBs may undergo serious degeneration at extreme pH values, leading to an increasingly disordered crimp and loose structure. An alkaline environment also contributes to the destruction of the phospholipid molecular structure (Zhao et al 2010), which leads to the decrease in G 00 of the OBs. The G 0 and G 00 values of three types of OBs at pH 4 were lower than that under the neutral condition, just as the Jms (2004) suggested.…”

Gel Electrophoresis of Protein - From Basic Science to Practical Approach

“…However, the precipitated proteins are difficult to re-dissolve. Incubation at 37 ºC, the use of a sample buffer with a high pH and high Tris concentration may be required for solubilisation (Zhao et al, 2010).…”