Strand and face: the topography of interactions between the SV40 origin of replication and T-antigen during the initiation of replication.

AUTOR(ES)

SenGupta, D J

RESUMO

The mechanism by which a replicator (origin of replication) becomes denatured during the initiation of replication is not understood for any prokaryotic or eukaryotic system. To address this question, we chemically probed the molecular contacts on the SV40 origin of replication (ori) that are used by the SV40 large T-antigen and a single-stranded DNA-binding protein (SSB) during ori denaturation. Prior to the actual denaturation step, the T-antigen double hexamer bound ori utilizing sugar-phosphate contacts that were located on opposite strands in each flanking domain of ori. Each set of flanking phosphate contacts were also located on approximately opposite faces of the ori duplex. While the phosphate contacts had a 2-fold symmetry with respect to the ori center, T-antigen contacts with nucleotide bases were polar with critical interactions detected in only one of the two flanking domains. During origin denaturation catalyzed by T-antigen and a SSB, numerous new contacts to flanking phosphates were observed on the strand not initially bound by T-antigen, suggesting movement of each T-antigen hexamer outward from ori. These data suggest that T-antigen initially binds ori in a manner that facilitates transfer of each T-antigen hexamer to opposite strands during the initiation of SV40 replication.

Documentos Relacionados

• Binding studies of SV40 T-antigen to SV40 binding site II.
• Simian virus 40 (SV40) DNA replication: SV40 large T antigen unwinds DNA containing the SV40 origin of replication.
• T-antigen binding to site I facilitates initiation of SV40 DNA replication but does not affect bidirectionality.
• Simian virus 40 (SV40) large tumor antigen causes stepwise changes in SV40 origin structure during initiation of DNA replication.
• Localized melting and structural changes in the SV40 origin of replication induced by T-antigen.