Herpes simplex virus DNA replication: a spacer sequence directs the ATP-dependent formation of a nucleoprotein complex at oriS.
The origin-binding protein (OBP) from herpes simplex virus 1 is a member of the SF2 helicase superfamily and is required for the initiation of DNA synthesis from a viral origin of DNA replication (oriS). The high-affinity binding sites for OBP in oriS, boxes I and II, are separated by an A+T-rich spacer. We used the gel retardation technique to examine the influence of this spacer sequence on the formation of a specific complex, referred to as complex II, between OBP and oriS. The formation of this OBP-oriS complex was greatly promoted by adenosine 5'-[gamma-thio]triphosphate and other nucleotide cofactors. Surprisingly, oriS constructs where the spacer sequence had been altered with approximately half of a helical turn (+4 or -6 base pairs) supported the formation of a more stable complex II than the wild-type origin. DNase I footprinting experiments showed that the cooperative binding of OBP to boxes I and II was affected by the length of the spacer sequence in the same way. In contrast, the ability of oriS-containing plasmids to replicate was most efficient with wild-type oriS. This paradox can be resolved if it is assumed that an ATP-dependent cooperative binding of OBP to properly spaced recognition sequences in oriS is required to induce a conformational change of DNA, thereby facilitating initiation of DNA replication.
Gustafsson
C M
CM
Department of Medical Biochemistry, Göteborg University, Sweden.
Hammarsten
O
O
Falkenberg
M
M
Elias
P
P
D000255:Adenosine Triphosphate / Q000378:metabolism*; D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D004261:DNA Replication; D004279:DNA, Viral / Q000096:biosynthesis*; D004268:DNA-Binding Proteins / Q000378:metabolism*; D003850:Deoxyribonuclease I; D008969:Molecular Sequence Data; D015394:Molecular Structure; D009036:Moths; D009690:Nucleic Acid Conformation; D009698:Nucleoproteins / Q000378:metabolism*; D011485:Protein Binding; D018139:Simplexvirus / Q000235:genetics*; D014764:Viral Proteins / Q000378:metabolism*
D004279:DNA, Viral; D004268:DNA-Binding Proteins; D009698:Nucleoproteins; D014764:Viral Proteins; C055789:origin-binding proteins, viral; D000255:Adenosine Triphosphate; D003850:Deoxyribonuclease I