title: Structural analysis of human Cdc20 supports multisite degron recognition by APC/C.
abstract: The anaphase-promoting complex/cyclosome (APC/C) promotes anaphase onset and mitotic exit through ubiquitinating securin and cyclin B1. The mitotic APC/C activator, the cell division cycle 20 (Cdc20) protein, directly interacts with APC/C degrons--the destruction (D) and KEN boxes. APC/C(Cdc20) is the target of the spindle checkpoint. Checkpoint inhibition of APC/C(Cdc20) requires the binding of a BubR1 KEN box to Cdc20. How APC/C recognizes substrates is not understood. We report the crystal structures of human Cdc20 alone or bound to a BubR1 KEN box. Cdc20 has a disordered N-terminal region and a C-terminal WD40 β propeller with a preformed KEN-box-binding site at its top face. We identify a second conserved surface at the side of the Cdc20 β propeller as a D-box-binding site. The D box of securin, but not its KEN box, is critical for securin ubiquitination by APC/C(Cdc20). Although both motifs contribute to securin ubiquitination by APC/C(Cdh1), securin mutants lacking either motif are efficiently ubiquitinated. Furthermore, D-box peptides diminish the ubiquitination of KEN-box substrates by APC/C(Cdh1), suggesting possible competition between the two motifs. Our results indicate the lack of strong positive cooperativity between the two degrons of securin. We propose that low-cooperativity, multisite target recognition enables APC/C to robustly ubiquitinate diverse substrates and helps to drive cell cycle oscillations.
authors.affiliation: Department of Pharmacology, University of Texas Southwestern Medical Center, Dallas, TX 75390, USA.
authors.forename: Wei
authors.initials: W
authors.lastname: Tian
authors.forename: Bing
authors.initials: B
authors.lastname: Li
authors.forename: Ross
authors.initials: R
authors.lastname: Warrington
authors.forename: Diana R
authors.initials: DR
authors.lastname: Tomchick
authors.forename: Hongtao
authors.initials: H
authors.lastname: Yu
authors.forename: Xuelian
authors.initials: X
authors.lastname: Luo
mesh_terms: D064173:Anaphase-Promoting Complex-Cyclosome; D001665:Binding Sites; D002352:Carrier Proteins / Q000737:chemistry / Q000378:metabolism; D064199:Cdc20 Proteins; D018797:Cell Cycle Proteins / Q000737:chemistry* / Q000378:metabolism*; D017124:Conserved Sequence; D006801:Humans; D008958:Models, Molecular; D016296:Mutagenesis / Q000235:genetics; D011485:Protein Binding; D017346:Protein Serine-Threonine Kinases / Q000737:chemistry / Q000378:metabolism; D017434:Protein Structure, Tertiary; D059748:Proteolysis; D013379:Substrate Specificity; D043743:Ubiquitin-Protein Ligase Complexes / Q000378:metabolism*; D054875:Ubiquitination
chemical_list: D002352:Carrier Proteins; D064199:Cdc20 Proteins; D018797:Cell Cycle Proteins; C577488:CDC20 protein, human; D043743:Ubiquitin-Protein Ligase Complexes; D064173:Anaphase-Promoting Complex-Cyclosome; C584277:BUB1 protein, human; C090464:Bub1 spindle checkpoint protein; D017346:Protein Serine-Threonine Kinases