Growing evidence points to the importance of colocalization of kinases and their substrates by cells to acheive specific biological effects. Such colocalization appears to be mediated by scaffolding or anchoring proteins which colocalize the appropriate kinases and substrates. One such protein in the A Kinase Anchoring Protein (AKAP) which colocalizes protein kinase A (PKA) and its substrate neurophysin. The object of this project is to develop a flow cytometric assay for the measurement of AKP-PKA interactions. Biotin tagged AKAP has been produced and will be attached to microspheres. PKA regulatory subunit will be fluorescently labeled and its binding to immobilized AKAP will be measured by flow cytometry.
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