Faculty Opinions recommendation of Role of the Orc6 protein in origin recognition complex-dependent DNA binding and replication in Drosophila melanogaster.

2007 ◽  
Author(s):  
David Gilbert
Keyword(s):  
Drosophila Melanogaster ◽  
Dna Binding ◽  
Origin Recognition Complex ◽  
Origin Recognition

scholarly journals Structural basis of DNA replication origin recognition by human Orc6 protein binding with DNA

2020 ◽  
Vol 48 (19) ◽  
pp. 11146-11161
Author(s):  
Naining Xu ◽  
Yingying You ◽  
Changdong Liu ◽  
Maxim Balasov ◽  
Lee Tung Lun ◽  
...  
Keyword(s):  
Dna Replication ◽  
Dna Binding ◽  
Solution Structure ◽  
Origin Recognition Complex ◽  
Structural Basis ◽  
Dna Replication Origin ◽  
Dna Alterations ◽  
Replication Initiator ◽  
Binding Subunit ◽  
Origin Recognition

Abstract The six-subunit origin recognition complex (ORC), a DNA replication initiator, defines the localization of the origins of replication in eukaryotes. The Orc6 subunit is the smallest and the least conserved among ORC subunits. It is required for DNA replication and essential for viability in all species. Orc6 in metazoans carries a structural homology with transcription factor TFIIB and can bind DNA on its own. Here, we report a solution structure of the full-length human Orc6 (HsOrc6) alone and in a complex with DNA. We further showed that human Orc6 is composed of three independent domains: N-terminal, middle and C-terminal (HsOrc6-N, HsOrc6-M and HsOrc6-C). We also identified a distinct DNA-binding domain of human Orc6, named as HsOrc6-DBD. The detailed analysis of the structure revealed novel amino acid clusters important for the interaction with DNA. Alterations of these amino acids abolish DNA-binding ability of Orc6 and result in reduced levels of DNA replication. We propose that Orc6 is a DNA-binding subunit of human/metazoan ORC and may play roles in targeting, positioning and assembling the functional ORC at the origins.

scholarly journals DrosophilaHeterochromatin Protein 1 (HP1)/Origin Recognition Complex (ORC) Protein Is Associated with HP1 and ORC and Functions in Heterochromatin-induced Silencing

2001 ◽  
Vol 12 (6) ◽  
pp. 1671-1685 ◽  
Author(s):  
Mohammed Momin Shareef ◽  
Chadwick King ◽  
Mona Damaj ◽  
RamaKrishna Badagu ◽  
Da Wei Huang ◽  
...  
Keyword(s):  
Dna Binding ◽  
Origin Recognition Complex ◽  
Polytene Chromosomes ◽  
Binding Activity ◽  
Hmg Proteins ◽  
Mitotic Chromosomes ◽  
Origin Firing ◽  
Dna Binding Activity ◽  
Origin Recognition

Heterochromatin protein 1 (HP1) is a conserved component of the highly compact chromatin of higher eukaryotic centromeres and telomeres. Cytogenetic experiments in Drosophila have shown that HP1 localization into this chromatin is perturbed in mutants for the origin recognition complex (ORC) 2 subunit. ORC has a multisubunit DNA-binding activity that binds origins of DNA replication where it is required for origin firing. The DNA-binding activity of ORC is also used in the recruitment of the Sir1 protein to silence nucleation sites flanking silent copies of the mating-type genes inSaccharomyces cerevisiae. A fraction of HP1 in the maternally loaded cytoplasm of the early Drosophilaembryo is associated with a multiprotein complex containingDrosophila melanogaster ORC subunits. This complex appears to be poised to function in heterochromatin assembly later in embryonic development. Here we report the identification of a novel component of this complex, the HP1/ORC-associated protein. This protein contains similarity to DNA sequence-specific HMG proteins and is shown to bind specific satellite sequences and the telomere-associated sequence in vitro. The protein is shown to have heterochromatic localization in both diploid interphase and mitotic chromosomes and polytene chromosomes. Moreover, the gene encoding HP1/ORC-associated protein was found to display reciprocal dose-dependent variegation modifier phenotypes, similar to those for mutants in HP1 and the ORC 2 subunit.

scholarly journals Architecture of the yeast origin recognition complex bound to origins of DNA replication.

1997 ◽  
Vol 17 (12) ◽  
pp. 7159-7168 ◽  
Author(s):  
D G Lee ◽  
S P Bell
Keyword(s):  
Dna Replication ◽  
Dna Binding ◽  
Origin Recognition Complex ◽  
Binding Assays ◽  
Chromosomal Replication ◽  
Origins Of Replication ◽  
Eukaryotic Dna ◽  
Level Of Analysis ◽  
Origin Recognition

In many organisms, the replication of DNA requires the binding of a protein called the initiator to DNA sites referred to as origins of replication. Analyses of multiple initiator proteins bound to their cognate origins have provided important insights into the mechanism by which DNA replication is initiated. To extend this level of analysis to the study of eukaryotic chromosomal replication, we have investigated the architecture of the Saccharomyces cerevisiae origin recognition complex (ORC) bound to yeast origins of replication. Determination of DNA residues important for ORC-origin association indicated that ORC interacts preferentially with one strand of the ARS1 origin of replication. DNA binding assays using ORC complexes lacking one of the six subunits demonstrated that the DNA binding domain of ORC requires the coordinate action of five of the six ORC subunits. Protein-DNA cross-linking studies suggested that recognition of origin sequences is mediated primarily by two different groups of ORC subunits that make sequence-specific contacts with two distinct regions of the DNA. Implications of these findings for ORC function and the mechanism of initiation of eukaryotic DNA replication are discussed.

scholarly journals Impacts of a new transcription factor family

2004 ◽  
Vol 166 (6) ◽  
pp. 765-768 ◽  
Author(s):  
Said Hashemolhosseini ◽  
Michael Wegner
Keyword(s):  
Transcription Factor ◽  
Drosophila Melanogaster ◽  
Dna Binding ◽  
Family Member ◽  
Parathyroid Gland ◽  
Dna Binding Domain ◽  
Transcription Factor Family ◽  
Master Regulators ◽  
Zinc Cations

GCM proteins constitute a small transcription factor family with a DNA-binding domain exhibiting a novel fold composed of two subdomains rigidly held together by coordination of one of two structural zinc cations. In all known cases, GCM proteins exert the role of master regulators: the prototypical family member determines gliogenesis in Drosophila melanogaster, whereas mammalian GCM proteins orchestrate divergent aspects of development and physiology in placenta, kidney, thymus, and parathyroid gland. Recent data point to an involvement of GCM proteins in different pathological contexts, such as preeclampsia, hyper- or hypoparathyroidism, and parathyroid gland tumors.

scholarly journals The role of ATP in the function of human ORC4 protein

2010 ◽  
Vol 75 (3) ◽  
pp. 317-322
Author(s):  
Aleksandra Divac ◽  
Branko Tomic ◽  
Jelena Kusic
Keyword(s):  
Conformational Changes ◽  
Origin Recognition Complex ◽  
Structural Changes ◽  
Atp Hydrolysis ◽  
Protein A ◽  
Structural Role ◽  
A Subunit ◽  
Dna Substrates ◽  
Origin Recognition

Human ORC4 protein, a subunit of the origin recognition complex, belongs to the AAA+ superfamily of ATPases. Proteins belonging to this family require ATP for their function and interactions with ATP lead to conformational changes in them or in their partners. Human ORC4 protein induces structural changes in DNA substrates, promoting renaturation and formation of non-canonical structures, as well as conversion of single-stranded into multi-stranded oligonucleotide structures. The aim of this study was to further investigate the role of ATP in the function of human ORC4 protein. For this purpose, a mutant in the conserved Walker B motif of ORC4, which is able to bind but not to hydrolyze ATP, was constructed and its activity in DNA restructuring reactions was investigated. The obtained results showed that ATP hydrolysis is not necessary for the function of human ORC4. It is proposed that ATP has a structural role as a cofactor in the function of human ORC4 as a DNA restructuring agent.

Analysis of mutant origin recognition complex with reduced ATPase activity in vivo and in vitro

2008 ◽  
Vol 413 (3) ◽  
pp. 535-543 ◽  
Author(s):  
Masaya Takehara ◽  
Masaki Makise ◽  
Hitomi Takenaka ◽  
Teita Asano ◽  
Tohru Mizushima
Keyword(s):  
Atpase Activity ◽  
Origin Recognition Complex ◽  
S Phase ◽  
Yeast Cells ◽  
Chromosomal Dna ◽  
Aaa Atpases ◽  
Origin Recognition

In eukaryotes, ORC (origin recognition complex), a six-protein complex, is the most likely initiator of chromosomal DNA replication. ORC belongs to the AAA+ (ATPases associated with a variety of cellular activities) family of proteins and has intrinsic ATPase activity derived from Orc1p, one of its subunits. To reveal the role of this ATPase activity in Saccharomyces cerevisiae (baker's yeast) ORC, we mutated the Orc1p sensor 1 and sensor 2 regions, which are important for ATPase activity in AAA+ proteins. Plasmid-shuffling analysis revealed that Asn600, Arg694 and Arg704 are essential for the function of Orc1p. In yeast cells, overexpression of Orc1R694Ep inhibited growth, caused inefficient loading of MCM (mini-chromosome maintenance complex of proteins) and slowed the progression of S phase. In vitro, purified ORC-1R [ORC with Orc1R694Ep (Orc1p Arg694→Glu mutant)] has decreased ATPase activity in the presence or absence of origin DNA. However, other activities (ATP binding and origin DNA binding) were indistinguishable from those of wild-type ORC. The present study showed that Arg694 of the Orc1p subunit is important for the ATPase activity of ORC and suggests that this ATPase activity is required for efficient MCM loading on to origin DNA and for progression of S phase.

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