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PMID: 16228006 Published · ppublish English Comparative Study Journal Article Research Support, N.I.H., Extramural Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, Non-P.H.S.

ATPase-dependent cooperative binding of ORC and Cdc6 to origin DNA.

Nature structural & molecular biology ·Vol. 12 ·No. 11 ·2005-11-00 ·Pages 965-71

Speck C, Chen Z, Li H, Stillman B

Abstract

Binding of Cdc6 to the origin recognition complex (ORC) is a key step in the assembly of a pre-replication complex (pre-RC) at origins of DNA replication. ORC recognizes specific origin DNA sequences in an ATP-dependent manner. Here we demonstrate cooperative binding of Saccharomyces cerevisiae Cdc6 to ORC on DNA in an ATP-dependent manner, which induces a change in the pattern of origin binding that requires the Orc1 ATPase. The reaction is blocked by specific origin mutations that do not interfere with the interaction between ORC and DNA. Single-particle reconstruction of electron microscopic images shows that the ORC-Cdc6 complex forms a ring-shaped structure with dimensions similar to those of the ring-shaped MCM helicase. The ORC-Cdc6 structure is predicted to contain six AAA+ subunits, analogous to other ATP-dependent protein machines. We suggest that Cdc6 and origin DNA activate a molecular switch in ORC that contributes to pre-RC assembly.

MeSH Terms
Amino Acid Sequence Cell Cycle Proteins/metabolism,ultrastructure DNA/genetics,metabolism DNA Footprinting DNA Primers Electrophoretic Mobility Shift Assay Microscopy, Electron Models, Molecular Molecular Sequence Data Multiprotein Complexes/chemistry,ultrastructure Mutation/genetics Origin Recognition Complex/metabolism,ultrastructure Saccharomyces cerevisiae Saccharomyces cerevisiae Proteins/metabolism,ultrastructure Structure-Activity Relationship
Chemicals
CDC6 protein, S cerevisiae Cell Cycle Proteins DNA Primers Multiprotein Complexes ORC1 protein, S cerevisiae Origin Recognition Complex Saccharomyces cerevisiae Proteins DNA
Authors & Affiliations
4 authors, click to expand affiliations / ORCID
Speck Christian
Cold Spring Harbor Laboratory, 1 Bungtown Road, Cold Spring Harbor, New York 11724, USA.
Chen Zhiqiang
Li Huilin
Stillman Bruce
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Article Info
Journal
Nature structural & molecular biology
Abbr.
Nat Struct Mol Biol
ISSN
1545-9993
DOI
10.1038/nsmb1002
Published
2005-11-00
Pages
965-71
Language
English
Region
United States
NLM ID
101186374
PMCID
PMC2952294
Subset
IM
Grants
NIGMS NIH HHS · R01 GM045436-20 · United States
NIGMS NIH HHS · R01 GM074985 · United States
NIGMS NIH HHS · R35 GM131754 · United States
NIGMS NIH HHS · GM45436 · United States
NIGMS NIH HHS · R01 GM045436 · United States
External Links
PubMed source DOI: 10.1038/nsmb1002 PMC Full Text View journal details
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