Click here to close
Hello! We notice that you are using Internet Explorer, which is not supported by Xenbase and may cause the site to display incorrectly.
We suggest using a current version of Chrome,
FireFox, or Safari.
???displayArticle.abstract???
The eukaryotic six-subunit origin recognition complex (ORC) governs the initiation site of DNA replication and formation of the prereplication complex. In this report we describe the isolation of the wild-type Homo sapiens (Hs)ORC and variants containing a Walker A motif mutation in the Orc1, Orc4, or Orc5 subunit using the baculovirus-expression system. Coexpression of all six HsORC subunits yielded a stable complex containing HsOrc subunits 1-5 (HsORC1-5) with virtually no Orc6 protein (Orc6p). We examined the ATPase, DNA-binding, and replication activities of these complexes. Similar to other eukaryotic ORCs, wild-type HsORC1-5 possesses ATPase activity that is stimulated only 2-fold by single-stranded DNA. HsORC1-5 with a mutated Walker A motif in Orc1p contains no ATPase activity, whereas a similar mutation of either the Orc4 or Orc5 subunit did not affect this activity. The DNA-binding activity of HsORC1-5, using lamin B2 DNA as substrate, is stimulated by ATP 3- to 5-fold. Mutations in the Walker A motif of Orc1p, Orc4p, or Orc5p reduced the binding efficiency of HsORC1-5 modestly (2- to 5-fold). Xenopus laevis ORC-depleted extracts supplemented with HsORC1-5 supported prereplication complex formation and X. laevis sperm DNA replication, whereas the complex with a mutation in the Walker A motif of the Orc1, Orc4, or Orc5 subunit did not. These studies indicate that the ATP-binding motifs of Orc1, Orc4, and Orc5 are all essential for the replication activity associated with HsORC.
Abdurashidova,
Localization of proteins bound to a replication origin of human DNA along the cell cycle.
2003, Pubmed
Abdurashidova,
Localization of proteins bound to a replication origin of human DNA along the cell cycle.
2003,
Pubmed Antunes,
Motifs in Schizosaccharomyces pombe ars3002 important for replication origin activity in Saccharomyces cerevisiae.
2003,
Pubmed Bell,
The origin recognition complex: from simple origins to complex functions.
2002,
Pubmed Bell,
ATP-dependent recognition of eukaryotic origins of DNA replication by a multiprotein complex.
1992,
Pubmed Bielinsky,
Origin recognition complex binding to a metazoan replication origin.
2001,
Pubmed Carpenter,
Role for a Xenopus Orc2-related protein in controlling DNA replication.
1996,
Pubmed
,
Xenbase Chesnokov,
A cytokinetic function of Drosophila ORC6 protein resides in a domain distinct from its replication activity.
2003,
Pubmed Chesnokov,
Assembly of functionally active Drosophila origin recognition complex from recombinant proteins.
1999,
Pubmed
,
Xenbase Chesnokov,
Functional analysis of mutant and wild-type Drosophila origin recognition complex.
2001,
Pubmed Chuang,
Purification and characterization of the Schizosaccharomyces pombe origin recognition complex: interaction with origin DNA and Cdc18 protein.
2002,
Pubmed Clyne,
Identification of autonomously replicating sequence (ARS) elements in eukaryotic cells.
1997,
Pubmed Dhar,
Architecture of the human origin recognition complex.
2001,
Pubmed Diffley,
DNA replication: building the perfect switch.
2001,
Pubmed Dutta,
Initiation of DNA replication in eukaryotic cells.
1997,
Pubmed Gillespie,
Reconstitution of licensed replication origins on Xenopus sperm nuclei using purified proteins.
2001,
Pubmed
,
Xenbase Kalejta,
Distal sequences, but not ori-beta/OBR-1, are essential for initiation of DNA replication in the Chinese hamster DHFR origin.
1998,
Pubmed Kelly,
Regulation of chromosome replication.
2000,
Pubmed Klemm,
Coordinate binding of ATP and origin DNA regulates the ATPase activity of the origin recognition complex.
1997,
Pubmed Kong,
Site-specific DNA binding of the Schizosaccharomyces pombe origin recognition complex is determined by the Orc4 subunit.
2001,
Pubmed Ladenburger,
Identification of a binding region for human origin recognition complex proteins 1 and 2 that coincides with an origin of DNA replication.
2002,
Pubmed Lee,
ATPase switches controlling DNA replication initiation.
2000,
Pubmed Lee,
Regulation of origin recognition complex conformation and ATPase activity: differential effects of single-stranded and double-stranded DNA binding.
2000,
Pubmed Lee,
The Schizosaccharomyces pombe origin recognition complex interacts with multiple AT-rich regions of the replication origin DNA by means of the AT-hook domains of the spOrc4 protein.
2001,
Pubmed Lee,
Architecture of the yeast origin recognition complex bound to origins of DNA replication.
1997,
Pubmed Loo,
The origin recognition complex in silencing, cell cycle progression, and DNA replication.
1995,
Pubmed Murray,
Cell cycle extracts.
1991,
Pubmed Rao,
The origin recognition complex interacts with a bipartite DNA binding site within yeast replicators.
1995,
Pubmed Remus,
DNA topology, not DNA sequence, is a critical determinant for Drosophila ORC-DNA binding.
2004,
Pubmed Siegel,
Determination of molecular weights and frictional ratios of proteins in impure systems by use of gel filtration and density gradient centrifugation. Application to crude preparations of sulfite and hydroxylamine reductases.
1966,
Pubmed Todorovic,
Replication origins of mammalian chromosomes: the happy few.
1999,
Pubmed Vashee,
Sequence-independent DNA binding and replication initiation by the human origin recognition complex.
2003,
Pubmed
,
Xenbase Vashee,
Assembly of the human origin recognition complex.
2001,
Pubmed
