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 metaphase plate. We report the solution structure of human Mad2 and its interaction with Cdc20. Mad2 possesses a novel three-layered alpha/beta fold with three alpha-helices packed between two beta-sheets. Using deletion mutants we identified the minimal Mad2-binding region of human Cdc20 as a 40-residue segment immediately N-terminal to the WD40 repeats. Mutagenesis and NMR titration experiments show that a C-terminal flexible region of Mad2 is required for binding to Cdc20. Mad2 and Cdc20 form a tight 1:1 heterodimeric complex in which the C-terminal segment of Mad2 becomes folded. These results provide the first structural insight into mechanisms of the spindle assembly checkpoint.
Luo
X
X
Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, 240 Longwood Avenue, Boston, Massachusetts 02115, USA.
Fang
G
G
Coldiron
M
M
Lin
Y
Y
Yu
H
H
Kirschner
M W
MW
Wagner
G
G
D000595:Amino Acid Sequence; D001665:Binding Sites; D002135:Calcium-Binding Proteins / Q000037:antagonists & inhibitors / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D002352:Carrier Proteins / Q000737:chemistry* / Q000378:metabolism*; D064199:Cdc20 Proteins; D018797:Cell Cycle Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D019281:Dimerization; D005656:Fungal Proteins / Q000037:antagonists & inhibitors / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D064246:Mad2 Proteins; D008958:Models, Molecular; D008969:Molecular Sequence Data; D019906:Nuclear Magnetic Resonance, Biomolecular; D009687:Nuclear Proteins; D017510:Protein Folding; D020836:Protein Structure, Quaternary; D017433:Protein Structure, Secondary; D012097:Repressor Proteins; D029701:Saccharomyces cerevisiae Proteins; D016415:Sequence Alignment; D017384:Sequence Deletion / Q000235:genetics; D012996:Solutions; D008941:Spindle Apparatus / Q000378:metabolism*; D013329:Structure-Activity Relationship
C577495:CDC20 protein, S cerevisiae; D002135:Calcium-Binding Proteins; D002352:Carrier Proteins; D064199:Cdc20 Proteins; D018797:Cell Cycle Proteins; D005656:Fungal Proteins; C577695:MAD2 protein, S cerevisiae; C577697:MAD2L1 protein, human; D064246:Mad2 Proteins; D009687:Nuclear Proteins; D012097:Repressor Proteins; D029701:Saccharomyces cerevisiae Proteins; D012996:Solutions