Abstract
Plant vacuole is a multifunctional organelle which storage solutes and proteins, digests them, control cytosolic pH and ions, fill large volume of cell space and others. Vacuole also contributes to dynamic events, such as Ca2+ signaling and stomatal movement. These functions were supported by a great number of tonoplast proteins and their interactions. In this study, we searched protein complex of vacuolar membrane comprehensively.
We prepared vacuolar membrane from suspension-cultured cell of Arabidopsis thaliana. Solubilized membranes were subjected to 2D- BN- SDS- PAGE to separate complexes of membrane protein. We detected approximately 20 candidates for hetero-oligomeric protein complex and about 20 candidates for homo-oligomeric protein. Using antibody library for vacuolar-membrane transporters of about 40, we analyzed apparent molecular masses of protein (complex) that contains target antigen under BN-PAGE. We also tried immunoprecipitation of the solubilized vacuolar membrane using the antibody library to pull down protein complexes of transporters.
