Sequence-independent DNA binding and replication initiation by the human origin recognition complex.
We report that a highly purified human origin recognition complex (HsORC) has intrinsic DNA-binding activity, and that this activity is modestly stimulated by ATP. HsORC binds preferentially to synthetic AT-rich polydeoxynucleotides, but does not effectively discriminate between natural DNA fragments that contain known human origins and control fragments. The complex fully restores DNA replication to ORC-depleted Xenopus egg extracts, providing strong evidence for its initiator function. Strikingly, HsORC stimulates initiation from any DNA sequence, and it does not preferentially replicate DNA containing human origin sequences. These data provide a biochemical explanation for the observation that in metazoans, initiation of DNA replication often occurs in a seemingly random pattern, and they have important implications for the nature of human origins of DNA replication.
Vashee
Sanjay
S
Institute for Biological Energy Alternatives, Rockville, Maryland 20850, USA.
Cvetic
Christin
C
Lu
Wenyan
W
Simancek
Pamela
P
Kelly
Thomas J
TJ
Walter
Johannes C
JC
D000818:Animals; D001667:Binding, Competitive; D002460:Cell Line; D002474:Cell-Free System; D004247:DNA / Q000378:metabolism*; D004261:DNA Replication; D004268:DNA-Binding Proteins; D004305:Dose-Response Relationship, Drug; D006801:Humans; D007313:Insecta; D051738:Origin Recognition Complex; D010957:Plasmids / Q000378:metabolism; D011485:Protein Binding; D011994:Recombinant Proteins / Q000378:metabolism; D013997:Time Factors; D014981:Xenopus
D004268:DNA-Binding Proteins; C497224:ORC2 protein, human; D051738:Origin Recognition Complex; D011994:Recombinant Proteins; D004247:DNA