Biochemical analysis of components of the pre-replication complex of Archaeoglobus fulgidus

被引:51
作者
Grainge, I [1 ]
Scaife, S [1 ]
Wigley, DB [1 ]
机构
[1] Canc Res UK, Clare Hall Labs, London Res Inst, Potters Bar EN6 3LD, Herts, England
关键词
D O I
10.1093/nar/gkg662
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
The eukaryotic pre-replication complex is assembled at replication origins in a reaction called licensing. Licensing involves the interactions of a variety of proteins including the origin recognition complex (ORC), Cdc6 and the Mcm2-7 helicase, homologues of which are also found in archaea. The euryarchaeote Archaeoglobus fulgidus encodes two genes with homology to Orc/Cdc6 and a single Mcm homologue. The A.fulgidus Mcm protein and one Orc/Cdc6 homologue have been purified and investigated in vitro. The Mcm protein is an ATP-dependent, hexameric helicase that can unwind between 200 and 400 bp of duplex DNA. Deletion of 112 amino acids from the N-terminus of A.f Mcm produced a protein, which was still capable of forming a hexamer, was competent in DNA binding and was able to unwind at least 1 kb of duplex DNA. The purified Orc/Cdc6 homologue was also able to bind DNA. Both Mcm and Orc/Cdc6 show a preference for specific DNA structures, namely molecules containing a single stranded bubble that mimics early replication intermediates. Nuclease protection showed that the binding sites for Mcm and Orc/Cdc6 overlap. The Orc/Cdc6 protein bound more tightly to these substrates and was able to displace pre-bound Mcm hexamer.
引用
收藏
页码:4888 / 4898
页数:11
相关论文
共 30 条
[1]   Components and dynamics of DNA replication complexes in S-cerevisiae: Redistribution of MCM proteins and Cdc45p during S phase [J].
Aparicio, OM ;
Weinstein, DM ;
Bell, SP .
CELL, 1997, 91 (01) :59-69
[2]   DNA replication in eukaryotic cells [J].
Bell, SP ;
Dutta, A .
ANNUAL REVIEW OF BIOCHEMISTRY, 2002, 71 :333-374
[3]   Archaea and the cell cycle [J].
Bernander, R .
MOLECULAR MICROBIOLOGY, 1998, 29 (04) :955-961
[4]   Mapping protein-protein interactions within a stable complex of DNA primase and DnaB helicase from Bacillus stearothermophilus [J].
Bird, LE ;
Pan, H ;
Soultanas, P ;
Wigley, DB .
BIOCHEMISTRY, 2000, 39 (01) :171-182
[5]   A double-hexamer archaeal minichromosome maintenance protein is an ATP-dependent DNA helicase [J].
Chong, JPJ ;
Hayashi, MK ;
Simon, MN ;
Xu, RM ;
Stillman, B .
PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA, 2000, 97 (04) :1530-1535
[6]   An integrated analysis of the genome of the hyperthermophilic archaeon Pyrococcus abyssi [J].
Cohen, GN ;
Barbe, V ;
Flament, D ;
Galperin, M ;
Heilig, R ;
Lecompte, O ;
Poch, O ;
Prieur, D ;
Quérellou, J ;
Ripp, R ;
Thierry, JC ;
Van der Oost, J ;
Weissenbach, J ;
Zivanovic, Y ;
Forterre, P .
MOLECULAR MICROBIOLOGY, 2003, 47 (06) :1495-1512
[7]   The Xenopus Cdc6 protein is essential for the initiation of a single round of DNA replication in cell-free extracts [J].
Coleman, TR ;
Carpenter, PB ;
Dunphy, WG .
CELL, 1996, 87 (01) :53-63
[8]   Archaea and the origin(s) of DNA replication proteins [J].
Edgell, DR ;
Doolittle, WF .
CELL, 1997, 89 (07) :995-998
[9]   Genome sequence of the hyperthermophilic crenarchaeon Pyrobaculum aerophilum [J].
Fitz-Gibbon, ST ;
Ladner, H ;
Kim, UJ ;
Stetter, KO ;
Simon, MI ;
Miller, JH .
PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA, 2002, 99 (02) :984-989
[10]   The structure and function of MCM from archaeal M-thermoautotrophicum [J].
Fletcher, RJ ;
Bishop, BE ;
Leon, RP ;
Sclafani, RA ;
Ogata, CM ;
Chen, XJS .
NATURE STRUCTURAL BIOLOGY, 2003, 10 (03) :160-167