Effect of extraction methods on the 2-DE map of whey proteome in cow milk

Abstract Text: For all proteomic approaches, protein extraction and sample preparation are the most critical steps for optimal results. However, protein isolation is often complicated. The objective of this study was to investigate the effect of extraction methods on two-dimensional electrophoresis (2-DE) maps of whey proteome in cow milk and explore an optimal eatraction method. Samples were extracted from cow milk by different isoelectric precipitations and ultracentrifugation methods. SDS-PAGE and 2-DE maps were analysised by QuantityOne v4.62 and PDQuest 8.0 software, respectively. The results of 2-DE maps showed that milk whey protein could be effectively extracted by the methods above with less background, no significant strips and good repeatability. However, there were still some residual caseins appearing in each map. 2-DE maps of whey protein refined by isoelectric precipitations were relatively informative compared with the gel maps obtained by ultracentrifugation. Moreover, the richness of different whey proteins in various maps extracted by the method adjusting pH to 4.6 as isoelectric point is slightly higher than adjusting pH to 4.8. The results indicated that adjusting pH to 4.6 as isoelectric point to extract whey protein has some advantages than the other two methods. However, all the methods used in this research could effectively remove high abundant casein to improve the detection sensitivity of low abundance proteins, which could provide some useful information for the futher research on whey protein proteomic of cow milk.

Keywords: whey protein, extraction methods, two-dimensional electrophoresis maps