Spacing is crucial for coordination of domain functions within the simian virus 40 core origin of replication.
The simian virus 40 core origin of replication is composed of distinct domains that are bracketed by DNA spacers. We created a matched set of insertion mutations in spacer sites to study the spatial relationships among origin domains. Insertions larger than a single base pair severely inhibit replication regardless of the helical phasing between domains. Replication-defective mutations reduce T-antigen binding and T-antigen-induced KMnO4 modifications of DNA to various extents. Mutations in the early half of the origin reduce T-antigen functions in the entire origin, whereas mutations in the late half reduce functions only in that half. Surprisingly, some mutations that severely inhibit DNA replication reduce T-antigen-induced melting and other structural changes within origin DNA to only a limited extent. In contrast, all replication-defective origin mutations prevent T antigen from extending the primary replication bubble beyond the limits of the core origin of replication. We conclude, therefore, that T-antigen-induced events within the core origin must be spatially coordinated for conversion of T-antigen hexamers bound to the core origin into mobile helicase units.
Parsons
R
R
Department of Microbiology, State University of New York, Stony Brook 11794-8621.
Tegtmeyer
P
P
D001483:Base Sequence; D004261:DNA Replication / Q000235:genetics*; D004279:DNA, Viral / Q000096:biosynthesis* / Q000235:genetics; D003850:Deoxyribonuclease I / Q000378:metabolism; D008969:Molecular Sequence Data; D016254:Mutagenesis, Insertional; D011196:Potassium Permanganate / Q000494:pharmacology; D013539:Simian virus 40 / Q000235:genetics* / Q000502:physiology; D014162:Transfection; D014779:Virus Replication / Q000235:genetics
D004279:DNA, Viral; D011196:Potassium Permanganate; D003850:Deoxyribonuclease I