March 28, 2024: Prof Wolfgang Peti, University of Connecticut Health Center

Описание: Using cryo-EM to understand dynamic interactions

As a trained NMR spectroscopist and experienced X-ray crystallographer, I will provide our newly gained experience and insights from using cryo-EM as a routine tool in our laboratory. As an example, I will use a recent complex that we determined by cryo-EM: PP2A:B55-ARPP19. PP2A:B55-ARPP19 is critical as progression through the cell cycle is controlled by regulated and abrupt changes in phosphorylation. Mitotic entry is initiated by increased phosphorylation of mitotic proteins, a process driven by kinases, while mitotic exit is achieved by counteracting dephosphorylation, a process driven by phosphatases, especially PP2A:B55.
While the role of kinases in mitotic entry is well-established, recent data have shown that mitosis is only successfully initiated when the counterbalancing phosphatases are also inhibited. For PP2A:B55, inhibition is achieved by the two intrinsically disordered proteins (IDPs), ARPP19 and FAM122A. Despite their critical roles in mitosis, the mechanisms by which they achieve PP2A:B55 inhibition is unknown. Here, we report the single particle cryo-electron microscopy (cryo-EM) structures of PP2A:B55 bound to phosphorylated ARPP19 and FAM122A. Consistent with our complementary NMR spectroscopy studies both IDPs bind PP2A:B55, but do so in highly distinct manners, unexpectedly leveraging multiple distinct binding sites on B55. Our extensive structural, biophysical and biochemical data explain how substrates and inhibitors are recruited to PP2A:B55 and provides a molecular roadmap for the development of therapeutic interventions for PP2A:B55 related diseases.