|Budget Amount *help
¥1,600,000 (Direct Cost : ¥1,600,000)
Fiscal Year 1998 : ¥700,000 (Direct Cost : ¥700,000)
Fiscal Year 1997 : ¥900,000 (Direct Cost : ¥900,000)
In the axon, many different kinds of membranous organelles are transported along microtubules.Kinesin super family-proteins (KIF), such as KIFlA, KIF1B, KIF2, KIR3A/B, KIF4, and KIF5, are the motor proteins for the anterograde transport of the membranous organelles.It is highly suggested that each KIF probably transports unique cargoes : KIFlA transports a subset of precursors of synaptic vesicles, and KIF1B transports the mitochondria.However, nothing is known about how the different KIFs recognize their specific cargoes.The homology of the sequences among different KIFs is high within the motor domain, but the sequences are rather varied outside the motor domain.It is known that KIFs bind to microtubules at the motor domain.It is likely that KIFs bind to the receptor on the membrane organelles at the regions outside the motor domain.To elucidate the mechanisms of binding of KIFs to the membrane organelles, we tried to identify and sequence the binding proteins for KIF3A/B/KAP3 complex on the membrane organelles.For this purpose, we used PC12 cell line, which are the good models for the neuronal cells and express abundant KIF3A/B/KAP3 complex, and identified the several possible candidates for the binding proteins using immunoprecipitation, metabolic labeling, and western blotting.To obtain the partial amino acid sequences, which are necessary for cloning and further characterization of these proteins, we developed the method for the large scale isolation of these proteins.We are now in the final stage of preparation of the samples for the determination of the partial amino acid sequences of the proteins.