Structure of the Mad2 spindle assembly checkpoint protein and its interaction with Cdc20

The checkpoint protein Mad2 inhibits the activity of the anaphase promoting complex by sequestering Cdc20 until all chromosomes are aligned at the met aphase plate. We report the solution structure of human Mad2 and its intera ction with Cdc20. Mad2 possesses a novel three-layered alpha/beta fold with three a-helices packed between two beta-sheets. Using deletion mutants we identified the minimal Mad2-binding region of human Cdc20 as a 40-residue s egment immediately N-terminal to the WD40 repeats. Mutagenesis and NMR titr ation experiments show that a C-terminal flexible region of Mad2 is require d for binding to Cdc20. Mad2 and Cdc20 form a tight 1:1 heterodimeric compl ex in which the C-terminal segment of Mad2 becomes folded. These results pr ovide the first structural insight into mechanisms of the spindle assembly checkpoint.