A dimerized coiled-coil domain and an adjoining part of geminin interact with two sites on Cdt1 for replication inhibition.
Geminin is a cellular protein that associates with Cdt1 and inhibits Mcm2-7 loading during S phase. It prevents multiple cycles of replication per cell cycle and prevents episome replication. It also directly inhibits the HoxA11 transcription factor. Here we report that geminin forms a parallel coiled-coil homodimer with atypical residues in the dimer interface. Point mutations that disrupt the dimerization abolish interaction with Cdt1 and inhibition of replication. An array of glutamic acid residues on the coiled-coil domain surface interacts with positive charges in the middle of Cdt1. An adjoining region interacts independently with the N-terminal 100 residues of Cdt1. Both interactions are essential for replication inhibition. The negative residues on the coiled-coil domain and a different part of geminin are also required for interaction with HoxA11. Therefore a rigid cylinder with negative surface charges is a critical component of a bipartite interaction interface between geminin and its cellular targets.
Saxena
Sandeep
S
Department of Biochemistry and Molecular Genetics, University of Virginia, Charlottesville 22908, USA.
Yuan
Ping
P
Dhar
Suman Kumar
SK
Senga
Takeshi
T
Takeda
David
D
Robinson
Howard
H
Kornbluth
Sally
S
Swaminathan
Kunchithapadam
K
Dutta
Anindya
A
D000595:Amino Acid Sequence; D000818:Animals; D018797:Cell Cycle Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D003460:Crystallization; D004261:DNA Replication; D004268:DNA-Binding Proteins / Q000378:metabolism*; D019281:Dimerization; D064248:Geminin; D018698:Glutamic Acid / Q000737:chemistry; D018398:Homeodomain Proteins / Q000378:metabolism; D006801:Humans; D007399:Interphase; D008969:Molecular Sequence Data; D017354:Point Mutation / Q000235:genetics*; D017434:Protein Structure, Tertiary; D016196:S Phase; D012645:Selenomethionine / Q000737:chemistry / Q000378:metabolism; D029867:Xenopus Proteins; D014982:Xenopus laevis / Q000378:metabolism
C467199:CDT1 protein, human; D018797:Cell Cycle Proteins; D004268:DNA-Binding Proteins; C577998:GMNN protein, Xenopus; C577995:GMNN protein, human; D064248:Geminin; C484298:HOXA11 protein, human; D018398:Homeodomain Proteins; D029867:Xenopus Proteins; D018698:Glutamic Acid; D012645:Selenomethionine