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Title: Crystal structure of the eukaryotic origin recognition complex

; ;  [1];  [2]
  1. (JHU-MED)
  2. (
Publication Date:
Research Org.:
Argonne National Lab. (ANL), Argonne, IL (United States). Advanced Photon Source (APS)
Sponsoring Org.:
OSTI Identifier:
Resource Type:
Journal Article
Resource Relation:
Journal Name: Nature; Journal Volume: 519; Journal Issue: 03, 2015
Country of Publication:
United States

Citation Formats

Bleichert, Franziska, Botchan, Michael R., Berger , James M., and UCB). Crystal structure of the eukaryotic origin recognition complex. United States: N. p., 2015. Web. doi:10.1038/nature14239.
Bleichert, Franziska, Botchan, Michael R., Berger , James M., & UCB). Crystal structure of the eukaryotic origin recognition complex. United States. doi:10.1038/nature14239.
Bleichert, Franziska, Botchan, Michael R., Berger , James M., and UCB). 2015. "Crystal structure of the eukaryotic origin recognition complex". United States. doi:10.1038/nature14239.
title = {Crystal structure of the eukaryotic origin recognition complex},
author = {Bleichert, Franziska and Botchan, Michael R. and Berger , James M. and UCB)},
abstractNote = {},
doi = {10.1038/nature14239},
journal = {Nature},
number = 03, 2015,
volume = 519,
place = {United States},
year = 2015,
month = 3
  • Binding of the Origin Recognition Complex (ORC) to origins of replication marks the first step in the initiation of replication of the genome in all eukaryotic cells. Here, we report the structure of the active form of human ORC determined by X-ray crystallography and cryo-electron microscopy. The complex is composed of an ORC1/4/5 motor module lobe in an organization reminiscent of the DNA polymerase clamp loader complexes. A second lobe contains the ORC2/3 subunits. The complex is organized as a double-layered shallow corkscrew, with the AAA+ and AAA+-like domains forming one layer, and the winged-helix domains (WHDs) forming a topmore » layer. CDC6 fits easily between ORC1 and ORC2, completing the ring and the DNA-binding channel, forming an additional ATP hydrolysis site. Analysis of the ATPase activity of the complex provides a basis for understanding ORC activity as well as molecular defects observed in Meier-Gorlin Syndrome mutations.« less
  • The ubiquitous class I basic helix-loop-helix (bHLH) factor E47 forms heterodimers with multiple tissue specific class II bHLH proteins to regulate distinct differentiation pathways. In order to define how class I- class II heterodimer partners are selected, we determined the crystal structure of the E47-NeuroD1-bHLH dimer in complex with the insulin promoter E-box sequence. Purification of the bHLH domain of E47-NeuroD1 indicates that E47 heterodimers are stable in solution. The interactions between E47 and NeuroD1 in the heterodimer are comparable to the interactions between E47 monomers in the homodimer, including hydrogen bonding, buried hydrophobic surface, and packing interactions. This ismore » consistent with a model in which E47-NeuroD1 heterodimers are favored due to the instability of NeuroD1 homodimers. Although E47?NeuroD1 is oriented uniquely on the E-box sequence (CATCTG) within the promoter of the insulin gene, no direct contacts are observed with the central base pairs within this E-box sequence. We propose that concerted domain motions allow E47 to form specific base contacts in solution. NeuroD1 is restrained from adopting the same base contacts by an additional phosphate backbone interaction by the neurogenic-specific residue His115. Orienting E47-NeuroD1 on promoters may foster protein?protein contacts essential to initiate transcription.« less
  • Ribonuclease HI (RNase H) is a member of the nucleotidyl-transferase superfamily and endo-nucleolytically cleaves the RNA portion in RNA/DNA hybrids and removes RNA primers from Okazaki fragments. The enzyme also binds RNA and DNA duplexes but is unable to cleave either. Three-dimensional structures of bacterial and human RNase H catalytic domains bound to RNA/DNA hybrids have revealed the basis for substrate recognition and the mechanism of cleavage. In order to visualize the enzyme's interactions with duplex DNA and to establish the structural differences that afford tighter binding to RNA/DNA hybrids relative to dsDNA, we have determined the crystal structure ofmore » Bacillus halodurans RNase H in complex with the B-form DNA duplex [d(CGCGAATTCGCG)]2. The structure demonstrates that the inability of the enzyme to cleave DNA is due to the deviating curvature of the DNA strand relative to the substrate RNA strand and the absence of Mg{sup 2+} at the active site. A subset of amino acids engaged in contacts to RNA 2{prime}-hydroxyl groups in the substrate complex instead bind to bridging or non-bridging phosphodiester oxygens in the complex with dsDNA. Qualitative comparison of the enzyme's interactions with the substrate and inhibitor duplexes is consistent with the reduced binding affinity for the latter and sheds light on determinants of RNase H binding and cleavage specificity.« less