Critical spatial requirement within the origin of simian virus 40 DNA replication.
We inserted a single base pair into the center of a 27-base-pair palindrome within the replication origin of simian virus 40. The mutation did not directly alter the symmetry of the palindrome or the protein-binding sequences within the palindrome. DNA binding studies showed that subunits of the simian virus 40 A protein (T antigen) bound to each of the four recognition pentanucleotides in the origin palindrome but did so with reduced affinity in comparison with wild-type origins. The mutant origin cloned in a plasmid DNA failed to replicate in COS cells. Thus, precise spatial interactions among subunits of A protein are necessary for stable origin binding and are crucial for subsequent steps in the initiation of DNA replication. Furthermore, any possible functional interactions of the simian virus 40 A protein with cellular DNA would require a great fidelity of protein binding arrangements to initiate cellular DNA replication.
D001483:Base Sequence; D004261:DNA Replication; D004279:DNA, Viral / Q000032:analysis; D003850:Deoxyribonuclease I; D004706:Endodeoxyribonucleases / Q000378:metabolism; D009154:Mutation; D013539:Simian virus 40 / Q000235:genetics*; D013463:Sulfuric Acid Esters / Q000494:pharmacology; D014779:Virus Replication
D004279:DNA, Viral; D013463:Sulfuric Acid Esters; D004706:Endodeoxyribonucleases; D003850:Deoxyribonuclease I; C007482:dimethyl sulfate