Poster: Seed biology

Abs # 710: Selection of an efficient method to characterize soybean seed proteins for 2D and MALDI-TOF analysis

Presenter:	Natarajan, Savithiry S, natarajs@ba.ars.usda.gov
Authors	Natarajan, Savithiry S (A)   Chenping, Chenping Chenping@w (B)   Thomas, J caperna@an (C)   Wesley, M wgarrett@a (D)
Affiliations:	(A): USDA-Agricultural Research Service, PSI (B): USDA-Agricultural Research Service, ANRI,Growth Biology Laboratory (C): USDA-Agricultural Research Service, ANRI, Biotechnology &Germplasm Laboratory (D): University of Maryland, Department of Natural Resouce Sciences &Landscape Architecture

Two-dimensional polyacrylamide gel electrophoresis (2D-PAGE) is one of the most powerful proteomics tools for separation and quantification of proteins from a complex mixture. Several methods of 2D analysis of plant and seed proteins have been reported. In this study, we compared four different protein extraction methods, including trichloroacetic acid (TCA)/acetone, phenol, and lysis buffer with and without thiourea to determine the method that yielded the best separated soybean seed proteins by 2D-PAGE. The modified TCA/acetone method showed higher protein resolution, and spot intensity than the other three methods. In addition, several non-abundant proteins were strongly detected in samples extracted with the TCA/acetone method as compared to the other methods. Matrix-assisted laser desorption/ionization time of flight (MALDI-TOF) analysis of trypsinized protein spots from 2D gels showed beta conglycinin with 3 subunits, glycinin with acidic and basic polypeptide chains, 2S albumin, globulin, dehydrin, trypsin inhibitor, and seed maturation proteins. These results indicate that the modified TCA/acetone method is an efficient and rapid method for 2D analysis of soybean seed proteins and is also suitable for MALDI-TOF analysis and identification of separated proteins.