Purification and identification of rice soluble glycoproteins

Abstract

The total proteins was extracted from rice leaves, and quantified by Coomassie Brilliant Blue G-250. We got four classifications from rice total proteins by using Ammonium Sulfate Precipitation, which was very important because we got easy way to separate and classify the total proteins. Also, it was useful for the purification of glycop roteins. The four classifications then purified by using ConA-Sepharose 4B Affinity Chromatography, the liquid of glycoproteins was collected by tubes in the same time of the appearances of absorption peaks. We even got some dry samples by using vacuum freezing and drying technology. Next, we done SDS-PAGE and determination respectively by Periodic AcidSchiff (PAS) staining and Coomassie Brilliant Blue (CBB) R-250 staining. In the gel of CBB staining, there are near 30 strips of different glycoproteins (some are glycopeptides) in the last three classifications; but the gel of PAS staining only showed 7 strips of different glycoproteins (some are glycopeptides) only because of the sensitivities of PAS much lower than CBB staining. The last step, we choose three glycoproteins showed on the gel of 50%--80% glycoproteins classification, all of them have higher contents than others. We further purified each of the three glycoproteins directly from the SDS-PAGE gels. They are proved purified glycoproteins or glycopeptides by CBB and PAS staining again. The three glycoproteins are named respectively as RG1, RG2 and RG3.